Antibodies targeting a complex comprising non-classical HLA-I and neoantigen and their methods of use

ABSTRACT

Provided herein are antibodies that selectively bind to complex comprising a non-classical HLA-I (e.g. HLA-E) and a neoantigen having variable heavy chain domains (VH), variable light chain domains (VL), and complementarity determining regions (CDRs) as disclosed herein, as well as methods and uses thereof.

CROSS-REFERENCE

This application claims the benefit of U.S. Provisional Application No.63/032,886, filed on Jun. 1, 2020, which is incorporated herein byreference in its entirety.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in ASCII format and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Aug. 31, 2020, isnamed ABX006_SL.txt and is 11,197 bytes in size.

SUMMARY

Disclosed herein, in certain embodiments, are monoclonal antibodies oran antigen-binding fragments thereof, comprising a light chain variabledomain (VL) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 7. Disclosed herein, incertain embodiments, are monoclonal antibodies or antigen-bindingfragments thereof, comprising a light chain variable domain (VL)comprising an amino acid sequence at least 90% identical to an aminoacid sequence set forth as SEQ ID NO: 7. Disclosed herein, in certainembodiments, are monoclonal antibodies or antigen-binding fragmentsthereof, comprising a light chain variable domain (VL) comprising anamino acid sequence at least 95% identical to an amino acid sequence setforth as SEQ ID NO: 7. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising alight chain variable domain (VL) comprising an amino acid sequence atleast 99% identical to an amino acid sequence set forth as SEQ ID NO: 7.Disclosed herein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a light chain variabledomain (VL) comprising an amino acid sequence 100% identical to an aminoacid sequence set forth as SEQ ID NO: 7. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof comprise aheavy chain variable domain (VH) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO: 8.In some embodiments, the monoclonal antibodies or antigen-bindingfragments thereof comprise a heavy chain variable domain (VH) comprisingan amino acid sequence at least 90% identical to an amino acid sequenceset forth as SEQ ID NO: 8. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof comprise a heavy chainvariable domain (VH) comprising an amino acid sequence at least 95%identical to an amino acid sequence set forth as SEQ ID NO: 8. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof comprise a heavy chain variable domain (VH) comprising an aminoacid sequence at least 99% identical to an amino acid sequence set forthas SEQ ID NO: 8. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise a heavy chain variable domain(VH) comprising an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 8. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof selectively bind to acomplex comprising an HLA-E and a neoantigen. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof do not have abinding affinity to the HLA-E alone. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof do not have a bindingaffinity to the neoantigen alone. In some embodiments, the neoantigen isexpressed by an antigen processing machinery (APM)-proficient cell. Insome embodiments, the neoantigen is expressed by a TAP1/2-proficientcell. In some embodiments, the neoantigen comprises, consistsessentially of, or consists of a sequence according to SEQ ID NO: 18(VMAPRTLFL). In some embodiments, the HLA-E is HLA-E*0101 or HLA-E*0103.In some embodiments, the antibodies selectively bind to the complexcomprising: (a) the HLA-E*0101 and the neoantigen; (b) the HLA-E*0103and the neoantigen; or (c) the HLA-E*0101 and the neoantigen, and theHLA-E*0103 and the neoantigen. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof are murine antibodies,chimeric antibodies, camelid antibodies, humanized antibodies, or humanantibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are TCR-like antibodies. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are multispecific antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof arebispecific antibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are bispecific T cell engagers (BiTE).In some embodiments, the BiTE binds to a CD3 protein associated with a Tcell receptor (TCR). In some embodiments, the BiTE further comprises alight chain variable domain (VL) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO:15. In some embodiments, the BiTE further comprises a heavy chainvariable domain (VH) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 16. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are multifunctional antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof antibodyfurther comprise a conjugated therapeutic moiety. In some embodiments,the selective binding of the antibodies to the complex comprising theHLA-E and the neoantigen induces an immune response in a cell. In someembodiments, the immune response comprises activation of (i) NK cells,(ii) T cells, or (iii) NK cells and T cells. In some embodiments, the Tcell is a CD4+ T cell or a CD8+ T cell. In some embodiments, the immuneresponse comprises activation of cytotoxic T cells (CTLs). In someembodiments, the cell is a cancer cell.

Disclosed herein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a heavy chain variabledomain (VH) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 8. Disclosed herein, incertain embodiments, are monoclonal antibodies or antigen-bindingfragments thereof, comprising a heavy chain variable domain (VH)comprising an amino acid sequence at least 90% identical to an aminoacid sequence set forth as SEQ ID NO: 8. Disclosed herein, in certainembodiments, are monoclonal antibodies or antigen-binding fragmentsthereof, comprising a heavy chain variable domain (VH) comprising anamino acid sequence at least 95% identical to an amino acid sequence setforth as SEQ ID NO: 8. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising aheavy chain variable domain (VH) comprising an amino acid sequence atleast 99% identical to an amino acid sequence set forth as SEQ ID NO: 8.Disclosed herein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a heavy chain variabledomain (VH) comprising an amino acid sequence 100% identical to an aminoacid sequence set forth as SEQ ID NO: 8. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof comprise alight chain variable domain (VL) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO: 7.In some embodiments, the monoclonal antibodies or antigen-bindingfragments thereof comprise a light chain variable domain (VL) comprisingan amino acid sequence at least 90% identical to an amino acid sequenceset forth as SEQ ID NO: 7. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof comprise a light chainvariable domain (VL) comprising an amino acid sequence at least 95%identical to an amino acid sequence set forth as SEQ ID NO: 7. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof comprise a light chain variable domain (VL) comprising an aminoacid sequence at least 99% identical to an amino acid sequence set forthas SEQ ID NO: 7. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise a light chain variable domain(VL) comprising an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 7. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof selectively binds to acomplex comprising an HLA-E and a neoantigen. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof do not have abinding affinity to the HLA-E alone. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof do not have a bindingaffinity to the neoantigen alone. In some embodiments, the neoantigen isexpressed by an antigen processing machinery (APM)-proficient cell. Insome embodiments, the neoantigen is expressed by a TAP1/2-proficientcell. In some embodiments, the neoantigen comprises, consistsessentially of, or consists of a sequence according to SEQ ID NO: 18(VMAPRTLFL). In some embodiments, the HLA-E is HLA-E*0101 or HLA-E*0103.In some embodiments, the antibodies selectively binds to the complexcomprising: (a) the HLA-E*0101 and the neoantigen; (b) the HLA-E*0103and the neoantigen; or (c) the HLA-E*0101 and the neoantigen, and theHLA-E*0103 and the neoantigen. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof are murine antibodies,chimeric antibodies, camelid antibodies, humanized antibodies, or humanantibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are TCR-like antibodies. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are multispecific antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof arebispecific antibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are bispecific T cell engagers (BiTE).In some embodiments, the BiTE binds to a CD3 protein associated with a Tcell receptor (TCR). In some embodiments, the BiTE further comprises alight chain variable domain (VL) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO:15. In some embodiments, the BiTE further comprises a heavy chainvariable domain (VH) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 16. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are multifunctional antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof antibodyfurther comprise a conjugated therapeutic moiety. In some embodiments,the selective binding of the antibody to the complex comprising theHLA-E and the neoantigen induces an immune response in a cell. In someembodiments, the immune response comprises activation of (i) NK cells,(ii) T cells, or (iii) NK cells and T cells. In some embodiments, the Tcell is a CD4+ T cell or a CD8+ T cell. In some embodiments, the immuneresponse comprises activation of cytotoxic T cells (CTLs). In someembodiments, the cell is a cancer cell.

Disclosed herein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising alight chain complementarity determining region (CDR) having an aminoacid sequence at least 90% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3. Disclosed herein, in certainembodiments, are monoclonal antibodies or antigen-binding fragmentsthereof, comprising a light chain complementarity determining region(CDR) having an amino acid sequence at least 95% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3. Disclosedherein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising alight chain complementarity determining region (CDR) having an aminoacid sequence 100% identical to at least one of the amino acid sequencesset forth as SEQ ID NOS: 1-3. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof comprise a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof comprise a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 90% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof comprise a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 95% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof comprise a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof comprise a heavy chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise a light chain variable domain(VL) comprising an amino acid sequence at least 80% identical to anamino acid sequence set forth as SEQ ID NO: 7. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof comprise aheavy chain variable domain (VH) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO: 8.In some embodiments, the monoclonal antibodies or antigen-bindingfragments thereof selectively binds to a complex comprising an HLA-E anda neoantigen. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof do not have a binding affinity to theHLA-E alone. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof do not have a binding affinity to theneoantigen alone. In some embodiments, the neoantigen is expressed by anantigen processing machinery (APM)-proficient cell. In some embodiments,the neoantigen is expressed by a TAP1/2-proficient cell. In someembodiments, the neoantigen comprises, consists essentially of, orconsists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL). In someembodiments, the HLA-E is HLA-E*0101 or HLA-E*0103. In some embodiments,the antibodies selectively binds to the complex comprising: theHLA-E*0101 and the neoantigen; the HLA-E*0103 and the neoantigen; or theHLA-E*0101 and the neoantigen, and the HLA-E*0103 and the neoantigen. Insome embodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are murine antibodies, chimeric antibodies, camelid antibodies,humanized antibodies, or human antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof are TCR-likeantibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are multispecific antibodies. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are bispecific antibodies. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof are bispecific T cellengagers (BiTE). In some embodiments, the BiTE binds to a CD3 proteinassociated with a T cell receptor (TCR). In some embodiments, the BiTEfurther comprises a light chain complementarity determining region (CDR)having an amino acid sequence at least 80% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 9-11. In someembodiments, the BiTE further comprises a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 80%identical to at least one of the amino acid sequences set forth as SEQID NOS: 12-14. In some embodiments, the BiTE further comprises a lightchain variable domain (VL) comprising an amino acid sequence at least80% identical to an amino acid sequence set forth as SEQ ID NO: 15. Insome embodiments, the BiTE further comprises a heavy chain variabledomain (VH) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 16. In some embodiments,the monoclonal antibodies or antigen-binding fragments thereof aremultifunctional antibodies. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof antibody furthercomprise a conjugated therapeutic moiety. In some embodiments, theselective binding of the antibody to the complex comprising the HLA-Eand the neoantigen induces an immune response in a cell. In someembodiments, the immune response comprises activation of (i) NK cells,(ii) T cells, or (iii) NK cells and T cells. In some embodiments, the Tcell is a CD4+ T cell or a CD8+ T cell. In some embodiments, the immuneresponse comprises activation of cytotoxic T cells (CTLs). In someembodiments, the cell is a cancer cell.

Disclosed herein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising aheavy chain complementarity determining region (CDR) having an aminoacid sequence at least 90% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 4-6. Disclosed herein, in certainembodiments, are monoclonal antibodies or antigen-binding fragmentsthereof, comprising a heavy chain complementarity determining region(CDR) having an amino acid sequence at least 95% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6. Disclosedherein, in certain embodiments, are monoclonal antibodies orantigen-binding fragments thereof, comprising a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. Disclosed herein, in certain embodiments, aremonoclonal antibodies or antigen-binding fragments thereof, comprising aheavy chain complementarity determining region (CDR) having an aminoacid sequence 100% identical to at least one of the amino acid sequencesset forth as SEQ ID NOS: 4-6. In some embodiments, the monoclonalantibody or antigen-binding fragment comprises a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3. In some embodiments, the monoclonal antibodyor antigen-binding fragment comprises a light chain complementaritydetermining region (CDR) having an amino acid sequence at least 90%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, the monoclonal antibody orantigen-binding fragment comprises a light chain complementaritydetermining region (CDR) having an amino acid sequence at least 95%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, the monoclonal antibody orantigen-binding fragment comprises a light chain complementaritydetermining region (CDR) having an amino acid sequence at least 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, the monoclonal antibody orantigen-binding fragment comprises a light chain complementaritydetermining region (CDR) having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 1-3.In some embodiments, the monoclonal antibodies or antigen-bindingfragments thereof comprise a heavy chain variable domain (VH) comprisingan amino acid sequence at least 80% identical to an amino acid sequenceset forth as SEQ ID NO: 8. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof comprise a light chainvariable domain (VL) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 7. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof selectively binds to a complex comprising an HLA-E and aneoantigen. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof do not have a binding affinity to theHLA-E alone. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof do not have a binding affinity to theneoantigen alone. In some embodiments, the neoantigen is expressed by anantigen processing machinery (APM)-proficient cell. In some embodiments,the neoantigen is expressed by a TAP1/2-proficient cell. In someembodiments, the neoantigen comprises, consists essentially of, orconsists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL). In someembodiments, the HLA-E is HLA-E*0101 or HLA-E*0103. In some embodiments,the antibodies selectively bind to the complex comprising: (a) theHLA-E*0101 and the neoantigen; (b) the HLA-E*0103 and the neoantigen; or(c) the HLA-E*0101 and the neoantigen, and the HLA-E*0103 and theneoantigen. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are murine antibodies, chimericantibodies, camelid antibodies, humanized antibodies, or humanantibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are TCR-like antibodies. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof are multispecific antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof arebispecific antibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are bispecific T cell engagers (BiTE).In some embodiments, the BiTE binds to a CD3 protein associated with a Tcell receptor (TCR). In some embodiments, the BiTE further comprises alight chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 9-11. In some embodiments, the BiTEfurther comprises a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 80% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 12-14. In someembodiments, the BiTE further comprises a light chain variable domain(VL) comprising an amino acid sequence at least 80% identical to anamino acid sequence set forth as SEQ ID NO: 15. In some embodiments, theBiTE further comprises a heavy chain variable domain (VH) comprising anamino acid sequence at least 80% identical to an amino acid sequence setforth as SEQ ID NO: 16. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof are multifunctional antibodies. Insome embodiments, the monoclonal antibodies or antigen-binding fragmentsthereof antibody further comprise a conjugated therapeutic moiety. Insome embodiments, the selective binding of the antibody to the complexcomprising the HLA-E and the neoantigen induces an immune response in acell. In some embodiments, the immune response comprises activation of(i) NK cells, (ii) T cells, or (iii) NK cells and T cells. In someembodiments, the cell is a CD4+ T cell or a CD8+ T cell. In someembodiments, the immune response comprises activation of cytotoxic Tcells (CTLs). In some embodiments, the cell is a cancer cell.

Disclosed herein, in certain embodiments, are bispecific antibodies oran antigen-binding fragment thereof, comprising: a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 80% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; anda light chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 9-11; or a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14. In some embodiments, the bispecificantibodies comprise: (a) a light chain variable domain (VL) comprisingan amino acid sequence at least 80% identical to an amino acid sequenceset forth as SEQ ID NO: 7; or a heavy chain variable domain (VH)comprising an amino acid sequence at least 80% identical to an aminoacid sequence set forth as SEQ ID NO: 8; and (b) a light chain variabledomain (VL) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 15; or a heavy chainvariable domain (VH) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 16.Disclosed herein, in certain embodiments, are bispecific antibodies oran antigen-binding fragment thereof, comprising: a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 80% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; anda light chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 9-11; and a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14. In some embodiments, the bispecificantibodies comprise: (a) a light chain variable domain (VL) comprisingan amino acid sequence at least 80% identical to an amino acid sequenceset forth as SEQ ID NO: 7; and a heavy chain variable domain (VH)comprising an amino acid sequence at least 80% identical to an aminoacid sequence set forth as SEQ ID NO: 8; and (b) a light chain variabledomain (VL) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 15; and a heavy chainvariable domain (VH) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 16. In someembodiments, the bispecific antibodies selectively bind to a complexcomprising an HLA-E and a neoantigen. In some embodiments, thebispecific antibodies do not have a binding affinity to the HLA-E alone.In some embodiments, the bispecific antibodies do not have a bindingaffinity to the neoantigen alone. In some embodiments, the neoantigen isexpressed by an antigen processing machinery (APM)-proficient cell. Insome embodiments, the neoantigen is expressed by a TAP1/2-proficientcell. In some embodiments, the neoantigen comprises, consistsessentially of, or consists of a sequence according to SEQ ID NO: 18(VMAPRTLFL). In some embodiments, the HLA-E is HLA-E*0101 or HLA-E*0103.In some embodiments, the antibodies selectively binds to the complexcomprising: the HLA-E*0101 and the neoantigen; the HLA-E*0103 and theneoantigen; or the HLA-E*0101 and the neoantigen, and the HLA-E*0103 andthe neoantigen. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are murine antibodies, chimericantibodies, camelid antibodies, humanized antibodies, or humanantibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are bispecific T cell engagers (BiTE).In some embodiments, the BiTE binds to a CD3 protein associated with a Tcell receptor (TCR). In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are multifunctional antibodies. Insome embodiments, the monoclonal antibodies or antigen-binding fragmentsthereof antibody further comprise a conjugated therapeutic moiety. Insome embodiments, the selective binding of the antibody to the complexcomprising the HLA-E and the neoantigen induces an immune response in acell. In some embodiments, the immune response comprises activation of(i) NK cells, (ii) T cells, or (iii) NK cells and T cells. In someembodiments, the T cell is a CD4+ T cell or a CD8+ T cell. In someembodiments, the immune response comprises activation of cytotoxic Tcells (CTLs). In some embodiments, the cell is a cancer cell.

Disclosed herein, in certain embodiments, are pharmaceuticalcompositions comprising: (a) a monoclonal antibody or an antigen-bindingfragment thereof disclosed herein or a bispecific antibody or anantigen-binding fragment thereof disclosed herein; and (b) apharmaceutically acceptable carrier or excipient.

Disclosed herein, in certain embodiments, are methods of treating cancerin an individual in need thereof, comprising administering to theindividual an effective amount of a monoclonal antibody or anantigen-binding fragment thereof comprising a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3. Disclosed herein, in certain embodiments, aremethods of treating cancer in an individual in need thereof, comprisingadministering to the individual an effective amount of a monoclonalantibody or an antigen-binding fragment thereof comprising a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof comprise (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and (b) a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 80%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 90% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and (b) a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 90%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 95% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and (b) a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 95%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and (b) a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise (a) a light chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 1-3; and (b) a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof comprise a light chain variable domain (VL) comprising an aminoacid sequence at least 80% identical to an amino acid sequence set forthas SEQ ID NO: 7. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof comprise a heavy chain variable domain(VH) comprising an amino acid sequence at least 80% identical to anamino acid sequence set forth as SEQ ID NO: 8. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof selectivelybinds to a complex comprising an HLA-E and a neoantigen. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof do not have a binding affinity to the HLA-E alone. In someembodiments, the monoclonal antibodies or antigen-binding fragmentsthereof do not have a binding affinity to the neoantigen alone. In someembodiments, the neoantigen is expressed by an antigen processingmachinery (APM)-proficient cell. In some embodiments, the neoantigen isexpressed by a TAP1/2-proficient cell. In some embodiments, theneoantigen comprises, consists essentially of, or consists of a sequenceaccording to SEQ ID NO: 18 (VMAPRTLFL). In some embodiments, the HLA-Eis HLA-E*0101 or HLA-E*0103. In some embodiments, the antibodiesselectively binds to the complex comprising: the HLA-E*0101 and theneoantigen; the HLA-E*0103 and the neoantigen; or the HLA-E*0101 and theneoantigen, and the HLA-E*0103 and the neoantigen. In some embodiments,the monoclonal antibodies or antigen-binding fragments thereof aremurine antibodies, chimeric antibodies, camelid antibodies, humanizedantibodies, or human antibodies. In some embodiments, the monoclonalantibodies or antigen-binding fragments thereof are TCR-like antibodies.In some embodiments, the monoclonal antibodies or antigen-bindingfragments thereof are multispecific antibodies. In some embodiments, themonoclonal antibodies or antigen-binding fragments thereof arebispecific antibodies. In some embodiments, the monoclonal antibodies orantigen-binding fragments thereof are bispecific T cell engagers (BiTE).In some embodiments, the BiTE binds to a CD3 protein associated with a Tcell receptor (TCR). In some embodiments, the BiTE further comprises alight chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 9-11. In some embodiments, the BiTEfurther comprises a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 80% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 12-14. In someembodiments, the BiTE further comprises a light chain variable domain(VL) comprising an amino acid sequence at least 80% identical to anamino acid sequence set forth as SEQ ID NO: 15. In some embodiments, theBiTE further comprises a heavy chain variable domain (VH) comprising anamino acid sequence at least 80% identical to an amino acid sequence setforth as SEQ ID NO: 16. In some embodiments, the monoclonal antibodiesor antigen-binding fragments thereof are multifunctional antibodies. Insome embodiments, the monoclonal antibodies or antigen-binding fragmentsthereof antibody further comprise a conjugated therapeutic moiety. Insome embodiments, the selective binding of the antibody to the complexcomprising the HLA-E and the neoantigen induces an immune response in acell. In some embodiments, the immune response comprises activation of(i) NK cells, (ii) T cells, or (iii) NK cells and T cells. In someembodiments, the T cell is a CD4+ T cell or a CD8+ T cell. In someembodiments, the immune response comprises activation of cytotoxic Tcells (CTLs). In some embodiments, the cancer is breast cancer, kidneycancer, lung cancer, ovarian cancer, colorectal cancer, pancreaticcancer, choriocarcinoma, non-small cell lung carcinoma (NSCLC), gastriccancer, cervical cancer, or head and neck cancer. In some embodiments,the cancer is a myeloima, leukemia or lymphoma. In some embodiments, thecancer is acute myeloid leukemia (AML), multiple myeloma, or amyelodysplastic syndrome. In some embodiments, the cancer is a B cellmalignancy. In some embodiments, the cancer is mantel cell lymphoma.

Disclosed herein, in certain embodiments, are methods of treating cancerin an individual in need thereof, comprising administering to theindividual an effective amount of a bispecific antibody or anantigen-binding fragment thereof, comprising: a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 80% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; anda light chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 9-11; or a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14. In some embodiments, the bispecificantibodies comprise: a light chain variable domain (VL) comprising anamino acid sequence at least 80% identical to an amino acid sequence setforth as SEQ ID NO: 7; or a heavy chain variable domain (VH) comprisingan amino acid sequence at least 80% identical to an amino acid sequenceset forth as SEQ ID NO: 8; and a light chain variable domain (VL)comprising an amino acid sequence at least 80% identical to an aminoacid sequence set forth as SEQ ID NO: 15; or a heavy chain variabledomain (VH) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 16. Disclosed herein, incertain embodiments, are methods of treating cancer in an individual inneed thereof, comprising administering to the individual an effectiveamount of a bispecific antibody or an antigen-binding fragment thereof,comprising: a light chain complementarity determining region (CDR)having an amino acid sequence at least 80% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 1-3; and a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 80% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6; and a light chain complementarity determiningregion (CDR) having an amino acid sequence at least 80% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 9-11; anda heavy chain complementarity determining region (CDR) having an aminoacid sequence at least 80% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 12-14. In some embodiments, thebispecific antibodies comprise: a light chain variable domain (VL)comprising an amino acid sequence at least 80% identical to an aminoacid sequence set forth as SEQ ID NO: 7; and a heavy chain variabledomain (VH) comprising an amino acid sequence at least 80% identical toan amino acid sequence set forth as SEQ ID NO: 8; and a light chainvariable domain (VL) comprising an amino acid sequence at least 80%identical to an amino acid sequence set forth as SEQ ID NO: 15; and aheavy chain variable domain (VH) comprising an amino acid sequence atleast 80% identical to an amino acid sequence set forth as SEQ ID NO:16. In some embodiments, the bispecific antibody or antigen-bindingfragment thereof selectively binds to a complex comprising an HLA-E anda neoantigen. In some embodiments, the bispecific antibody orantigen-binding fragment thereof does not have a binding affinity to theHLA-E alone. In some embodiments, the bispecific antibody orantigen-binding fragment thereof does not have a binding affinity to theneoantigen alone. In some embodiments, the neoantigen is expressed by anantigen processing machinery (APM)-proficient cell. In some embodiments,the neoantigen is expressed by a TAP1/2-proficient cell. In someembodiments, the neoantigen comprises, consists essentially of, orconsists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL). In someembodiments, the HLA-E is HLA-E*0101 or HLA-E*0103. In some embodiments,the bispecific antibodies selectively bind to the complex comprising:the HLA-E*0101 and the neoantigen; the HLA-E*0103 and the neoantigen; orthe HLA-E*0101 and the neoantigen, and the HLA-E*0103 and theneoantigen. In some embodiments, the bispecific antibody orantigen-binding fragment thereof is a murine antibody, a chimericantibody, a camelid antibody, a humanized antibody, or a human antibody.In some embodiments, the bispecific antibody or antigen-binding fragmentthereof is a TCR-like antibody. In some embodiments, the bispecificantibody or antigen-binding fragment thereof is a bispecific T cellengager (BiTE). In some embodiments, the BiTE binds to a CD3 proteinassociated with a T cell receptor (TCR). In some embodiments, thebispecific antibody or antigen-binding fragment thereof is amultifunctional antibody. In some embodiments, the bispecific antibodyor antigen-binding fragment thereof antibody further comprise aconjugated therapeutic moiety. In some embodiments, the selectivebinding of the antibody to the complex comprising the HLA-E and theneoantigen induces an immune response in a cell. In some embodiments,the immune response comprises activation of (i) NK cells, (ii) T cells,or (iii) NK cells and T cells. In some embodiments, the T cell is a CD4+T cell or a CD8+ T cell. In some embodiments, the immune responsecomprises activation of cytotoxic T cells (CTLs). In some embodiments,the bispecific antibody is administered at a therapeutically effectiveamount. In some embodiments, the cancer is breast cancer, kidney cancer,lung cancer, ovarian cancer, colorectal cancer, pancreatic cancer,choriocarcinoma, non-small cell lung carcinoma (NSCLC), gastric cancer,cervical cancer, or head and neck cancer. In some embodiments, thecancer is a myeloima, leukemia or lymphoma. In some embodiments, thecancer is acute myeloid leukemia (AML), multiple myeloma, or amyelodysplastic syndrome. In some embodiments, the cancer is a B cellmalignancy. In some embodiments, the cancer is mantel cell lymphoma.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

FIG. 1A-FIG. 1D exemplifies identification and affinity characterizationof clone R4 clone 1 (R4c1) against V-0025. (FIG. 1A) Phage and (FIG. 1B)monoclonal soluble ELISA. Antigen is at 1 μg/ml. (FIG. 1C) Bindingaffinity sensorgram and (FIG. 1D) kinetic data.

FIG. 2A-FIG. 2D are exemplary overview of sorting schematic for theisolation of affinity matured clones from R4c1 FIG. 2A-FIG. 2C exemplifycell sorting plots showing each round of sorting for the first round ofaffinity maturation. The ‘star’ indicates the sorted population. For theR4 sort (FIG. 2D), Koff was utilized with V-0025 at 10 nM usingR4c1-IgG1 at 1 μM for the various timepoints as the antibody sink.

FIG. 3A-FIG. 3G exemplify clones identified from affinity maturation ofR4c1. FIG. 3A-FIG. 3D exemplify binding profile to target antigen(V-0025) at 100 nM. FIG. 3E-FIG. 3G exemplify binding profile of theclone, R2A_1, to target (V-0025) at 10 nM and to additionalpeptide/HLA-E antigens at 100 nM.

FIG. 4 exemplifies R2A_1 binding to A549-B4 cells.

FIG. 5A-FIG. 5B exemplify isolation process for improved clones byaffinity maturation of CDRL3 region of R2A_1. FIG. 5A exemplifiesbinding to target (V-0025) by parent clone (R4 clone 1) and the startingaffinity maturation library (RO) and first round sorted library (R1).FIG. 5B exemplifies Koff of R1 affinity maturation library. Target(V-0025) is at 10 nM, with R2A_1-hIgG1 as antibody sink at 1 μM for 15minutes up to 90 minutes.

FIG. 6A-FIG. 6B exemplify the sorting schematic for the isolation ofclones with greater binding affinity than the parent. Cell sorting plotsshowing (FIG. 6A) round 1 and (FIG. 6B) round 2 sorting for affinitymaturation. The asterisk indicates the sorted population. For the R2sort, Koff was utilized with V-0025 at 10 nM using R2A_1-IgG1 at 1 μMfor 45 minutes as the antibody sink.

FIG. 7 exemplifies identification of clones from affinity maturation ofthe CDRL3 region of R2A_1 and binding profile to V-0025 at 1 nM and 0.1nM. Arrow indicates the top clone.

FIG. 8A-FIG. 8B exemplify QC data for the production of ABX0011.

FIG. 9A-FIG. 9K exemplify binding specificity, affinity and stability ofABX0011. Specificity to antigen by ELISA (FIG. 9A) HLA-E (clone 3D12)and (FIG. 9B) ABX0011. For ELISA assays, antibodies were used at 1 mg/mland HLA-E/peptide antigens were used at 0.5 mg/ml. Monovalent affinityof ABX0011 to target V-0025 was determined using ResoSens label-freeinstrument (Resonant Sensors, Inc. Arlington, Tex.) (FIG. 9C). The limitof detection for ABX0011 compared to clone 3D12 was assessed by ELISA(FIGS. 9D&E). ABX0011 and HLA-E were used at 1 μg/mL while V-0025 wastitrated from 1 μg/mL down to 0.001 μg/mL (Fig. (D). FIG. 9E showstarget V-0025 used at 0.25 μg/mL while ABX0011 and HLA-E were titratedfrom 2 μg/mL down to 3.8 pg/mL (3.8×10⁻⁶ μg/mL). Thermostability ofABX0011 (FIG. 9F) was assessed by ELISA. ABX0011 binding toHLA-E/VMAPRTLFL (SEQ ID NO: 18) was evaluated after antibody wasincubated in mouse serum at 37° C. for 7 to 14 days. Selectivity ofbinding (FIG. 9G) of clone 3D12 and ABX0011 to unpulsed andpeptide-pulsed K562.E cells. K562.E cells were pulsed with 20 mM ofpeptide for 2 hrs followed by staining with antibody at 1 μg/mL. Furtherassessment of ABX0011 binding selectivity was performed using ABX0012(ABX0011 with mouse IgG1) with related and unrelated peptides. HLA-E(FIG. 9H) and ABX0012 (FIG. 9I) staining on peptide-pulsed and unpulsedK562.E cells. The limit of detection for ABX0011 was determined withtarget V-0025 peptide pulsed K562.E (FIG. 9J). Controls were unpulsedK562.E cells and parent K562 (lack HLA-E expression) and K562.E cellspulsed with irrelevant peptides (V-0034, V-0044, V-0046 and P-0550).Epitope mapping for ABX0011 was carried out using K562.E cells pulsedwith alanine substituted V-0025 peptides (FIG. 9K).

FIG. 10A-FIG. 10C exemplify ABX0011 binding specificity to a cell linethat expresses HLA-E and HLA-G (FIG. 10A) and no binding was observed tohuman peripheral blood mononuclear cells (huPBMC) and primary humanumbilical vascular endothelial cells (huVEC) lacking HLA-G (FIG. 10B andFIG. 10C). Antibody binding studies were performed at 1 μg/mL. (FIG.10A) Binding to IFNγ-stimulated JEG3 wild type (WT, top panel), E^(ko)(middle panel) and Tap-1^(ko) (bottom panel). (FIG. 10B) Binding tohuPBMC and (FIG. 10C) to huVEC.

FIG. 11A-FIG. 11B exemplify binding to AML cell lines that express bothHLA-E and HLA-G (THP-1 and OCI-AML3) or only express HLA-E (KG-1). AMLcells stained at 1 μg/ml of antibody using 3D12 (HLA-E), MEM-G/09(HLA-G) and W632 (pan HLA I) (FIG. 11A). ABX0012 (ABX0011 with mouseIgG1) was titrated (10 to 0.004 mg/ml) and binding to target positiveAML cell lines was observed (FIG. 11B).

FIG. 12A-FIG. 12D exemplify ABX0011 mediated NK cytotoxicity againsttumor cell lines expressing target. NK cells were co-cultured withV-0025 peptide-pulsed K562.E cells or with THP-1 cells with an isotypecontrol (hIgG1). The frequency of CD107a-positive NK cells andCD107a-positive NKG2A+ NK cells are shown (FIG. 12 A and FIG. 12B). Thefrequency of dead target cells (K562.E) and THP-1 cells is shown in FIG.12C and FIG. 12D, respectively.

FIG. 13A-FIG. 13B exemplify QC data for the production of ABX0030, abispecific T cell engager that contains the VL and VH domains of ABX0011linked covalently to an anti-CD3 scFv fragment.

FIG. 14 exemplifies affinity analysis for ABX0030.

FIG. 15 exemplifies that ABX0030 binding to K562.E cells is peptidespecific. Only K562.E cells pulsed with V-0025 target peptide werestained with ABX0030 in a dose dependent manner. Control cells, parentK562 and K562.E without peptide were not stained with ABX0030.

FIG. 16 are exemplary histograms to illustrate that ABX0030 stains humanCD3+ cells and not CD3neg cells. PBMCs stained with GAH-APC conjugateand anti-CD3 (clone SK7) (bottom left histogram). PBMCs stained withanti-CD4 and GAH-APC conjugate (upper left histogram). CD3+ cellstaining with ABX0030+GAH-APC conjugate (no SK7 Ab) (upper righthistogram). Double staining with anti-CD-4-PE and ABX0030+GAH-APCconjugate (upper right histogram). Notice in this panel the twopopulations of CD3+ cells representing CD4+ and CD8+ T-cells. Doublingstaining of CD3+ cells using ABX0030 and clone SK7-PE (bottom righthistogram).

FIG. 17A-FIG. 17C exemplify ABX0030 performance to specifically activateCD8+ T-cells and to redirect T-cell lysis of target positive cells. FIG.17A shows specific redirected T-cell lysis of K562.E cells pulsed withV-0025 target peptide by ABX0030. Some lysis activity was observed forK562.E (unpulsed) cells when ABX0030 was used at 10,000 pM (highestconcentration). CD8+ T cell CD25 and CD107a expression was alsoquantified to determine whether ABX0030 could non-specifically activateT cells. (FIG. 17B and FIG. 17C) ABX0030 exhibited minimal activation ofCD8+ T-cells in the presence of K562.E unpulsed cells (no target).

FIG. 18A-FIG. 18E exemplify potent activity of ABX0030 to redirect CD8+T-cell lysis of THP-1 cells. FIG. 18A shows a dose-dependent effect(10000 to 80 pM) of ABX0030 to redirect CD8+ T-cell lysis of THP-1cells. ABX0030 activated CD8+ T-cells in presence of THP-1 cells.

FIG. 18B shows ABX0030 dose-dependent effect on the frequency ofCD25+CD8+ T-cells. Additional markers demonstrating ABX0030 effect onCD8+ T-cells in presence of THP-1 cells are an increase in frequency ofCD107+CD8+ T-cells (FIG. 18C), and increases in the frequency ofperforin (FIG. 18D) and IFNg (FIG. 18E) positive CD8+ T-cells.

DETAILED DESCRIPTION

Disclosed herein, in certain embodiments, are antibodies comprising atleast one heavy chain comprising a heavy chain variable domain (VH) andat least one light chain comprising a light chain variable domain (VL).Each VH and VL comprises three complementarity determining regions(CDR). In some embodiments, the antibodies are bispecific antibodies. Insome embodiments, the bispecific antibodies are bispecific T cellengagers (BiTEs). In some embodiments, the bispecific antibodies bind toa CD3 protein associated with a T cell receptor (TCR). Further disclosedherein, in certain embodiments, are methods of treating a cancer byadministering an antibody that selectively binds to a complex comprisinga non-classical HLA-I (e.g. HLA-E) and a neoantigen. In someembodiments, the antibodies that selectively bind to a complexcomprising a non-classical HLA-I (e.g. HLA-E) and a neoantigen modulateimmune response against cancer cells, thereby treating cancer. In someembodiments, the antibodies are bispecific antibodies.

Traditional approaches to the treatment of cancers have includedsurgery, radiation, chemotherapy and hormone therapy. However, suchtherapies have not proven effective by themselves. Development ofalternate remedies for preventing and/or treating cancer is crucial.More recently immunotherapy and gene therapy approaches utilizingantibodies and T-lymphocytes have emerged as new and promising methodsfor treating cancer.

Major histocompatibility complex (MHC) molecules, designated humanleukocyte antigen (HLA) in humans, play a critical role in the body'srecognition of disease and the resulting immune response to cancer andinvading antigens. The HLA gene family is divided into two subgroupsnamely HLA Class I (HLA-I) and HLA Class II (HLA-II), with HLA-I furtherdivided into classical HLA-I and non-classical HLA-I. Each HLA moleculeforms a complex with one peptide from within the cell. On cancer cells,some of the peptide/HLA complexes are uniquely presented which enablesthe immune system to recognize and kill these cells. Cells decoratedwith these unique peptide/HLA complexes are recognized and killed by thecytotoxic T cells (CTLs). Cancer cells show a downregulation inclassical HLA-I expression but an upregulation in non-classical HLA-Iexpression (e.g. HLA-E). Thus, the upregulated uniquely presentednon-classical HLA-I-peptide complexes on cancer cells are novel targetsfor developing innovative immunotherapies for treatment of cancer.

Certain Terminology

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which the claimed subject matter belongs. It is to be understoodthat the foregoing general description and the following detaileddescription are exemplary and explanatory only and are not restrictiveof any subject matter claimed. The section headings used herein are fororganizational purposes only and are not to be construed as limiting thesubject matter described.

As used herein, singular forms “a”, “and,” and “the” include pluralreferents unless the context clearly indicates otherwise. Thus, forexample, reference to “an antibody” includes a plurality of antibodiesand reference to “an antibody” in some embodiments includes multipleantibodies, and so forth.

As used herein, all numerical values or numerical ranges include wholeintegers within or encompassing such ranges and fractions of the valuesor the integers within or encompassing ranges unless the context clearlyindicates otherwise. Thus, for example, reference to a range of 90-100%,includes 91%, 92%, 93%, 94%, 95%, 95%, 97%, etc., as well as 91.1%,91.2%, 91.3%, 91.4%, 91.5%, etc., 92.1%, 92.2%, 92.3%, 92.4%, 92.5%,etc., and so forth. In another example, reference to a range of 1-5,000fold includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,18, 19, 20, fold, etc., as well as 1.1, 1.2, 1.3, 1.4, 1.5, fold, etc.,2.1, 2.2, 2.3, 2.4, 2.5, fold, etc., and so forth.

“About” a number, as used herein, refers to range including the numberand ranging from 10% below that number to 10% above that number. “About”a range refers to 10% below the lower limit of the range, spanning to10% above the upper limit of the range.

As used herein, the term “MHC” refers to the Major HistocompabilityComplex, which is a set of gene loci specifying major histocompatibilityantigens. The term “HLA” as used herein refer to Human LeukocyteAntigens, which are the histocompatibility antigens found in humans. Asused herein, “HLA” is the human form of “MHC” and the terms are usedinterchangeably.

As used herein “antibody” refers to a glycoprotein which exhibitsbinding specificity to a specific antigen. Antibodies herein alsoinclude “antigen binding portion” or fragments of the antibody that arecapable of binding to the antigen. The term includes, but is not limitedto, polyclonal, monoclonal, monospecific, multispecific (e.g.,bispecific antibodies), natural, humanized, human, chimeric, synthetic,recombinant, hybrid, mutated, grafted, antibody fragments (e.g., aportion of a full-length antibody, generally the antigen binding orvariable region thereof, e.g., Fab, Fab′, F(ab′)2, and Fv fragments),and in vitro generated antibodies so long as they exhibit the desiredbiological activity. The term also includes single chain antibodies,e.g., single chain Fv (sFv or scFv) antibodies, in which a variableheavy and a variable light chain are joined together (directly orthrough a peptide linker) to form a continuous polypeptide.

As used herein “CDR” refers to an immunoglobulin (Ig) hypervariabledomain. A CDR is defined by any suitable manner. In some embodiments,CDRs can be defined in accordance with any of the Chothia numberingschemes, the Kabat numbering scheme, a combination of Kabat and Chothia,the AbM definition, the contact definition, and/or a combination of theKabat, Chothia, AbM, and/or contact definitions; and may producedifferent results.

As used herein, the term “selectively binds” in the context of anybinding agent, e.g., an antibody, refers to a binding agent that bindsspecifically to an antigen or epitope, such as with a high affinity, anddoes not significantly bind other unrelated antigens or epitopes.

As used herein the term “neoantigen” or “neopeptide” are usedinterchangeably and refer to a peptide differentially expressed by adiseased or stressed cell (e.g. cancer cell) compared to a healthy cell.

The terms “recipient”, “individual”, “subject”, “host”, and “patient”,are used interchangeably herein and in some cases, refer to anymammalian subject for whom diagnosis, treatment, or therapy is desired,particularly humans. None of these terms require the supervision ofmedical personnel.

As used herein, the terms “treatment,” “treating,” and the like, in somecases, refer to administering an agent, or carrying out a procedure, forthe purposes of obtaining an effect. The effect may be prophylactic interms of completely or partially preventing a disease or symptom thereofand/or may be therapeutic in terms of effecting a partial or completecure for a disease and/or symptoms of the disease. “Treatment,” as usedherein, may include treatment of a disease or disorder (e.g. cancer) ina mammal, particularly in a human, and includes: (a) preventing thedisease or a symptom of a disease from occurring in a subject which maybe predisposed to the disease but has not yet been diagnosed as havingit (e.g., including diseases that may be associated with or caused by aprimary disease; (b) inhibiting the disease, i.e., arresting itsdevelopment; and (c) relieving the disease, i.e., causing regression ofthe disease. Treating may refer to any indicia of success in thetreatment or amelioration or prevention of a cancer, including anyobjective or subjective parameter such as abatement; remission;diminishing of symptoms or making the disease condition more tolerableto the patient; slowing in the rate of degeneration or decline; ormaking the final point of degeneration less debilitating. The treatmentor amelioration of symptoms is based on one or more objective orsubjective parameters; including the results of an examination by aphysician. Accordingly, the term “treating” includes the administrationof the compounds or agents of the present invention to prevent or delay,to alleviate, or to arrest or inhibit development of the symptoms orconditions associated with diseases (e.g. cancer). The term “therapeuticeffect” refers to the reduction, elimination, or prevention of thedisease, symptoms of the disease, or side effects of the disease in thesubject.

A “therapeutically effective amount” in some cases means the amountthat, when administered to a subject for treating a disease, issufficient to effect treatment for that disease.

“Percent (%) identity” refers to the extent to which two sequences(nucleotide or amino acid) have the same residue at the same positionsin an alignment. For example, “an amino acid sequence is X % identicalto SEQ ID NO: Y” refers to % identity of the amino acid sequence to SEQID NO: Y and is elaborated as X % of residues in the amino acid sequenceare identical to the residues of sequence disclosed in SEQ ID NO: Y.Generally, computer programs are employed for such calculations.Exemplary programs that compare and align pairs of sequences, includeALIGN (Myers and Miller, 1988), FASTA (Pearson and Lipman, 1988;Pearson, 1990) and gapped BLAST (Altschul et al., 1997), BLASTP, BLASTN,or GCG (Devereux et al., 1984).

Major Histocompatibility Complex (MHC) or Human Leukocyte Antigens (HLA)

Major histocompatibility complexes (MHC), also termed Human LeukocyteAntigens (HLA) in humans are glycoproteins expressed on the surface ofnucleated cells that act as proteomic scanning chips by providinginsight into the status of cellular health. They continuously samplepeptides from normal host cellular proteins, cancer cells, inflamedcells and bacterial, viral and parasite infected cells and present shortpeptides on the surface of cells for recognition by T lymphocytes.Presented peptides can also be derived from proteins that are out offrame or from sequences embedded in the introns, or from proteins whosetranslation is initiated at codons other than the conventionalmethionine codon, ATG.

There are two classes of MHCs in mice and humans, namely MHC I and MHCII. WIC I comprises classical and non-classical MHC I sub-groups.

Classical MHC I or HLA-I

Classical WIC I molecules include HLA-A, HLA-B and HLA-C in humans andH-2-K, H-2-D, H-2-B and H-2-L in mice. Classical MHC I molecules arehighly polymorphic with more than 2,735 alleles of HLA-A, 3,455 allelesof HLA-B and 2,259 alleles of HLA-C. Classical MHC I is expressed on thesurface of all nucleated cells and present peptides to CD8 Tlymphocytes. 30% of the proteins in the cellular machinery are rapidlydegraded and are primary substrates for classical WIC I antigenpresentation.

For peptide to be presented by classical MHC I molecules, proteins arefirst processed through the conventional processing route (ubiquitinproteasome system) which begins with protein degradation in theproteasome and Transporter associated protein (TAP) dependent transportof peptides into the endoplasmic reticulum (ER) and ends with theloading of peptides into the HLA peptide binding pocket. The proteinsthat contribute to the conventional processing route are collectivelyknown as antigen processing machinery (APM) and include the proteasome,TAP complex, tapasin, endoplasmic reticulum amino peptidase (ERAAP),binding immunoglobulin protein (BiP), clanexin and calreticulin. Cellslacking either proteasome subunits, TAP1/2, ErP57 or calreticulin havereduced numbers of classical MHC I molecules on their surface.

Non-Classical MHC I or HLA-I

Non-classical MHC I molecules include HLA-E, HLA-F and HLA-G, and havelimited polymorphisms. They play a role in regulating innate andadaptive immune responses. Non-classical MHC I molecules presentpeptides generated by both the conventional processing route and thealternative processing route in health and disease states, and representa novel set of markers for targeting in disease states (e.g. cancer).

HLA-E

The non-classical MHC class I molecule, HLA-E is non-polymorphic. Innature, 13 HLA-E alleles have been identified with only two functionalvariants, namely HLAE* 0101 and HLA-E*0103. The difference betweenHLA-E*0101 (HLA-E^(107R)) and *0103 (HLA-E^(107G)) is a single aminoacid difference at position 107 which is outside the peptide bindingpocket. Similar to the classical MHC I molecules, HLA-E is expressed inall cells with a nucleus, however at usually lower levels. HLA-Emolecule expression in cells and tissues is generally increased duringstress and disease. As such, HLA-E is differentially expressed onstressed or diseased cell (e.g. cancer cell) compared to on a healthycell.

In healthy cells, HLA-E presents peptides derived from classical MHCmolecules and the non-classical HLA-G molecule to either inhibit orstimulate the activity of NK cells and a subset of CD8 T cells throughengaging the receptor CD94/NKG2. Depending on the particular peptidepresented by HLA-E, the HLA-E complex engages either CD94/NKG2A orCD94/NKG2C to inhibit or activate NK cells and a subset of CD8 T cells,respectively.

Another signal peptide that has characteristics in common with signalpeptides generated from classical HLA-I molecules is the signal peptidegenerated from non-classical HLA-G. HLA-G expression under normalphysiologic conditions is tightly regulated, with limited expressionfound in relatively few tissues and cells in the body. HLA-G plays a keyrole as an immune tolerant molecule and its expression is observed incancer tissue/cells. Moreover, the signal peptide from HLA-G isprocessed by the conventional antigen processing pathway and deliveredto the endoplasmic reticulum by the peptide transporter TAP. In someembodiments, the signal peptide is VMAPRTLFL (SEQ ID NO: 18).

HLA-E Expression and Peptide Presentation in Cancer Cells

Cells deficient in one or more components of the APM load peptides intoMHC class I molecules via alternative processing routes which areindependent of the APM-dependent conventional processing route.APM-deficient cells not only have reduced numbers of classical MHC Imolecules on their surface, but also show an increase in the cellsurface density of HLA-E molecules as well as an increase in therepertoire of peptides presented. The alternative processing routes areconstitutively turned on and produce peptides in both healthy anddiseased cells. These peptides, however, are not presented by healthycells; instead they are only presented in diseased or stressed cells. Assuch, the different peptide repertoires generated by APM-defectivecells, also known as “T-cell epitopes associated with impaired peptideprocessing” (TEIPP), represent novel targets unique to cancer cells, andrepresent ideal targets for therapeutic development in the treatment ofcancer.

In a stressed or diseased state (e.g. cancer), the stressed or diseasedcell (e.g. a cancer cell) differentially expresses a complex comprisingan HLA-E and a peptide derived from the non-classical HLA-G molecule.The complex comprising an HLA-E and a peptide derived from thenon-classical HLA-G molecule is differentially expressed on the stressedor diseased cell compared to a healthy cell. Targeting this complexinduces an immune response against the cell expressing the complex.

MHC II or HLA-H

MHC II molecules in humans include HLA-DM, HLA-DO, HLA-DP, HLA-DQ andHLA-DR and include H-2 I-A and H-2 I-E in mice. MHC II expression ismore restricted to B cells, dendritic cells, macrophages, activated Tcells and thymic epithelial cells and MHC II molecules present peptidesto CD4 lymphocytes.

Antibodies that Target a Complex Comprising a Non-Classical HLA-I (e.g.HLA-E) and a Neoantigen

Disclosed herein, in certain embodiments, are antibodies that target acomplex comprising a non-classical HLA-I (e.g. HLA-E) and a neoantigen.In some embodiments, the antibodies comprise at least one heavy chaincomprising a heavy chain variable domain (VH) and at least one lightchain comprising a light chain variable domain (VL). Each VH and VLcomprises three complementarity determining regions (CDR). The aminoacid sequences of the VH and VL and the CDRs determine the antigenbinding specificity and antigen binding strength of the antibody. Theamino acid sequences of the VH and VL and the CDRs are summarized inTable 1.

TABLE 1 Antibodies Human monoclonal antibody sequences that bind a  complex comprising HLA-E and classical HLA signal peptides (ABX-0010, 0011, 0030) SEQ ID SEQUENCE NO: Light Chain CDR1 QSISSY 1Light Chain CDR2 AAS 2 Light Chain CDR3 QQAAAYPSL 3 Heavy Chain CDR1GFTFSSYA 4 Heavy Chain CDR2 IAYGGGAT 5 Heavy Chain CDR3 AKGLSNFDY 6Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQ 7 Variable domainKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISS LQPEDFATYYCQQAAAYPSLFGQGTKVEIKHeavy Chain EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWV 8 Variable domainRQAPGKGLEWVSTIAYGGGATAYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKGLSNFDYWGQG TLVTVSSMouse monoclonal antibody sequences binding CD3 Light Chain CDR1TGAVTTSNY 9 Light Chain CDR2 GTN 10 Light Chain CDR3 ALWYSNLWV 11Heavy Chain CDR1 GFTFNTYA 12 Heavy Chain CDR2 IRSKYNNYAT 13Heavy Chain CDR3 VRHGNFNSYVSWFAYG 14 Light ChainQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWV 15 Variable domainQEKPDHLF TGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL Heavy ChainEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYANINW 16 Variable domainVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYV SWFAYWGQGTLVTVSS

In some embodiments, the antibodies selectively bind to a complexcomprising a non-classical HLA-I (e.g. HLA-E) and a neoantigen. In someembodiments, the antibody does not have a binding affinity to thenon-classical HLA-I alone. In some embodiments, the antibody does nothave a binding affinity to the neoantigen alone. In some embodiments,the antibody does not have a binding affinity to a complex comprisingthe non-classical HLA-I and a non-relevant neoantigen.

In some embodiments, the neoantigen is expressed by an antigenprocessing machinery (APM)-proficient cell. In some embodiments, theneoantigen is expressed by a TAP1/2-proficient cell. In someembodiments, the neoantigen comprises, consists essentially of, orconsists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL).

In some embodiments, the non-classical HLA-I is HLA-E, HLA-F, HLA-G, orHLA-H. In some embodiments, the non-classical HLA-I is HLA-E. In someembodiments, the HLA-E is HLA-E*0101. In some embodiments, the HLA-E isHLA-E*0103.

In some embodiments, the antibody selectively binds to the complexcomprising the HLA-E and the neoantigen. In some embodiments, theantibody selectively binds to the complex comprising the HLA-E*0101 andthe neoantigen. In some embodiments, the antibody selectively binds tothe complex comprising the HLA-E*0103 and the neoantigen. In someembodiments, the antibody selectively binds to the complex comprisingthe HLA-E*0101 and the neoantigen, and to the complex of the HLA-E*0103and the neoantigen. In some embodiments, the complex comprises the HLA-Eand a neoantigen according to SEQ ID NO: 18 (VMAPRTLFL).

In some embodiments, the antibody is a murine antibody. In someembodiments, the antibody is a chimeric antibody. In some embodiments,the antibody is a camelid antibody. In some embodiments, the antibody isa humanized antibody. In some embodiments, the antibody is a humanantibody.

In some embodiments, the antibody is a TCR-like antibody. In someembodiments, the antibody is a single domain antibody. In someembodiments, the single domain antibody is a camelid single domainantibody.

In some embodiments, the antibody is a multispecific antibody. In someembodiments, the antibody is a bispecific antibody. In some embodiments,the antibody is a bispecific T cell engager (BiTE). In some embodiments,the BiTE binds to a CD3 protein associated with a T cell receptor (TCR).In some embodiments, the BiTE binds to a CD3c protein associated with aT cell receptor (TCR). In some embodiments, the antibody is amultifunctional antibody.

In some embodiments, the antibody further comprises a conjugatedtherapeutic moiety. Therapeutic moiety include, but are not limited to,a cytotoxin, a chemotherapeutic drug, an immunosuppressant, and aradioisotope. A cytotoxin or cytotoxic agent includes any agent that isdetrimental to (e.g., kills) cells. Examples include, but are notlimited to, taxol, cytochalasin B, gramicidin D, ethidium bromide,emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine,colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione,mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone,glucocorticoids, procaine, tetracaine, lidocaine, propranolol, andpuromycin and analogs or homologs thereof. Suitable chemotherapeuticagents include, but are not limited to, anti metabolites (e.g.,methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, fludarabin,5-fluorouracil, decarbazine, hydroxyurea, azathiprin, gemcitabin andcladribin), alkylating agents (e.g., mechlorethamine, thioepa,chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU),cyclophosphamide, busulfan, dibromomannitol, streptozotocin, mitomycinC, and cis-dichlorodiamine platinum (II) (DDP) cisplatin),anthracyclines (e.g., daunorubicin (formerly daunomycin) anddoxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin),bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents(e.g., vincristine, vinblastine, docetaxel, paclitaxel and vinorelbin).Suitable radioisotopes include, but are not limited to, iodine-131,yttrium-90 or indium-Ill. Further examples of therapeutic moieties are aprotein or polypeptide possessing a desired biological activity. Suchproteins may include, for example, an enzymatically active toxin, oractive fragment thereof, such as abrin, ricin A, pseudomonas exotoxin,or diphtheria toxin; a protein such as tumor necrosis factor orinterferon-γ; or biological response modifiers such as, for example,lymphokines, interleukin-1 (IL-1), interleukin-2 (IL-2), interleukin-6(IL-6), granulocyte macrophage colony stimulating factor (GM-CSF),granulocyte colony stimulating factor (G-CSF), or other growth factors.

In some embodiments, the selective binding of the antibody to thecomplex comprising the non-classical HLA-I (e.g. HLA-E) and theneoantigen induces an immune response. In some embodiments, the immuneresponse comprises activation of NK cells. In some embodiments, theselective binding of the antibody to the CD3 protein associated with aTCR induces an immune response. In some embodiments, the immune responsecomprises activation of T cells. In some embodiments, the T cell is aCD8+ T cell. In some embodiments, the immune response comprisesactivation of cytotoxic T cells (CTLs). In some embodiments, the cell isa cancer cell.

Antibody Variable Domain (VL and VH)

Disclosed herein are antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen, the antibodies having a lightchain comprising a light chain variable domain (VL). In someembodiments, antibodies comprise a light chain variable domain (VL)having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 7. In some embodiments the VL hasan amino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 7. In someembodiments, the VL has an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 7.

Further disclosed herein are antibodies that selectively bind to acomplex comprising an HLA-E and a neoantigen, the antibodies having aheavy chain comprising a heavy chain variable domain (VH). In someembodiments, antibodies comprise a heavy chain variable domain (VH)having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 8. In some embodiments the VH hasan amino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 8. In someembodiments, the VH has an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 8.

Also disclosed herein are antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen, the antibodies comprising a lightchain variable domain (VL) and a heavy chain variable domain (VH). Insome embodiments, antibodies comprise a light chain variable domain (VL)having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 7 and a heavy chain variabledomain (VH) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 8. In some embodimentsthe VL has an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 7 and the VH has an amino acid sequence at least about 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence set forthas SEQ ID NO: 8. In some embodiments, the VL has an amino acid sequence100% identical to an amino acid sequence set forth as SEQ ID NO: 7 andthe VH has an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 8.

Antibody Complementarity Determining Regions (CDR)

Disclosed herein are antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen, the antibodies having a lightchain comprising a light chain complementarity determining region (CDR).In some embodiments, antibodies comprise a light chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3. In someembodiments, antibodies comprise a light chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, antibodies comprise a light chain CDRsequence having an amino acid sequence 100% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 1-3.

Further disclosed herein are antibodies that selectively bind to acomplex comprising an HLA-E and a neoantigen, the antibodies having aheavy chain comprising a heavy chain complementarity determining region(CDR). In some embodiments, antibodies comprise a heavy chain CDRsequence having an amino acid sequence at least about 70% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 4-6.In some embodiments, antibodies comprise a heavy chain CDR sequencehaving an amino acid sequence at least about 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, antibodies comprise aheavy chain CDR sequence having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 4-6.

Also disclosed herein are antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen comprise a light chaincomplementarity determining region (CDR) and a heavy chaincomplementarity determining region (CDR). In some embodiments,antibodies comprise a light chain CDR sequence having an amino acidsequence at least about 70% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3 and a heavy chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6. In someembodiments, antibodies comprise a light chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3 and a heavy chain CDR sequence having an amino acid sequenceat least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6. Insome embodiments, antibodies comprise a light chain CDR sequence havingan amino acid sequence 100% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3 and a heavy chain CDR sequencehaving an amino acid sequence at least about 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6.

In some embodiments, antibodies selectively bind to a complex comprisingan HLA-E and a neoantigen comprise at least one of a light chain CDR1having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 1, a light chain CDR2 having anamino acid sequence at least about 70% identical to an amino acidsequence set forth as SEQ ID NO: 2, and a light chain CDR3 having anamino acid sequence at least about 70% identical to an amino acidsequence set forth as SEQ ID NO: 3. In some embodiments, antibodiescomprise at least one of a light chain a light chain CDR1 having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 1, a lightchain CDR2 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 2, and a light chain CDR3 having an amino acid sequence atleast about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an aminoacid sequence set forth as SEQ ID NO: 3. In some embodiments, antibodiescomprise at least one of a light chain CDR1 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 1, a light chain CDR2 having an amino acid sequence 100% identicalto an amino acid sequence set forth as SEQ ID NO: 2, and a light chainCDR3 having an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 3.

In some embodiments, antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen comprise at least one of a heavychain CDR1 having an amino acid sequence at least about 70% identical toan amino acid sequence set forth as SEQ ID NO: 4, a heavy chain CDR2having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 5, a heavy chain CDR3 having anamino acid sequence at least about 70% identical to an amino acidsequence set forth as SEQ ID NO: 6. In some embodiments, antibodiescomprise at least one of a heavy chain CDR1 having an amino acidsequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%,88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identicalto an amino acid sequence set forth as SEQ ID NO: 4, a heavy chain CDR2having an amino acid sequence at least about 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:5, a heavy chain CDR3 having an amino acid sequence at least about 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 6. In some embodiments, antibodies comprise at leastone of a heavy chain CDR1 having an amino acid sequence 100% identicalto an amino acid sequence set forth as SEQ ID NO: 4, a heavy chain CDR2having an amino acid sequence 100% identical to an amino acid sequenceset forth as SEQ ID NO: 5, a heavy chain CDR3 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 6.

In some embodiments, antibodies that selectively bind to a complexcomprising an HLA-E and a neoantigen comprise at least one of a lightchain CDR1 having an amino acid sequence at least about 70% identical toan amino acid sequence set forth as SEQ ID NO: 1, a light chain CDR2having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 2, a light chain CDR3 having anamino acid sequence at least about 70% identical to an amino acidsequence set forth as SEQ ID NO: 3, a heavy chain CDR1 having an aminoacid sequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 5, and a heavy chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 6.In some embodiments, antibodies comprise at least one of a light chainCDR1 having an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 1, a light chain CDR2 having an amino acid sequence at least about75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 8′7%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 2, a light chain CDR3 having an amino acidsequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%,88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identicalto an amino acid sequence set forth as SEQ ID NO: 3, a heavy chain CDR1having an amino acid sequence at least about 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:4, a heavy chain CDR2 having an amino acid sequence at least about 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 5, and a heavy chain CDR3 having an amino acidsequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%,88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identicalto an amino acid sequence set forth as SEQ ID NO: 6. In someembodiments, antibodies comprise at least one of a light chain CDR1having an amino acid sequence 100% identical to an amino acid sequenceset forth as SEQ ID NO: 1, a light chain CDR2 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 2, a light chain CDR3 having an amino acid sequence 100% identicalto an amino acid sequence set forth as SEQ ID NO: 3, a heavy chain CDR1having an amino acid sequence 100% identical to an amino acid sequenceset forth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 5, and a heavy chain CDR3 having an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 6.

Bispecific Antibodies

In some embodiments, the antibodies disclosed herein are bispecificantibodies. In some embodiments, the bispecific antibodies arebispecific T cell engagers (BiTEs). In some embodiments, the BiTE bindsto a CD3 protein associated with a T cell receptor (TCR). In someembodiments, the BiTE binds to a CD3c protein associated with a T cellreceptor (TCR).

In some embodiments, the BiTE comprises an anti-CD3 antibody, or afragment thereof, such as UCHT1, OKT3, F6A, L2K, muromonab,otelixizumab, teplizumab, visilizumab, CD3-12, MEM-57, 4D10A6, CD3D, orTR66.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence at least70% identical to an amino acid sequence set forth as SEQ ID NO: 7; or aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; or a heavy chain variable domain (VH) comprising an aminoacid sequence at least 70% identical to an amino acid sequence set forthas SEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence at least75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 7; or a heavy chain variable domain (VH)comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to an amino acid sequence set forth as SEQ ID NO: 8; and(b) a light chain variable domain (VL) comprising an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an aminoacid sequence set forth as SEQ ID NO: 15; or a heavy chain variabledomain (VH) comprising an amino acid sequence at least 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 7; or aheavy chain variable domain (VH) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 15; or aheavy chain variable domain (VH) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence at least70% identical to an amino acid sequence set forth as SEQ ID NO: 7; and aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; and a heavy chain variable domain (VH) comprising anamino acid sequence at least 70% identical to an amino acid sequence setforth as SEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence at least75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 7; and a heavy chain variable domain (VH)comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to an amino acid sequence set forth as SEQ ID NO: 8; and(b) a light chain variable domain (VL) comprising an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an aminoacid sequence set forth as SEQ ID NO: 15; and a heavy chain variabledomain (VH) comprising an amino acid sequence at least 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain variable domain (VL) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 7; and aheavy chain variable domain (VH) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 15; and aheavy chain variable domain (VH) comprising an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence at least 70% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3; or a heavy chain complementaritydetermining region (CDR) having an amino acid sequence at least 70%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6; and (b) a light chain complementarity determining region(CDR) having an amino acid sequence at least 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 9-11; or aheavy chain complementarity determining region (CDR) having an aminoacid sequence at least 70% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 1-3; or aheavy chain complementarity determining region (CDR) having an aminoacid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%,89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; or a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) alight chain complementarity determining region (CDR) having an aminoacid sequence 100% identical to at least one of the amino acid sequencesset forth as SEQ ID NOS: 9-11; or a heavy chain complementaritydetermining region (CDR) having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence at least 70% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3; and a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6; and (b) a light chain complementaritydetermining region (CDR) having an amino acid sequence at least 70%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; and a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 70% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 1-3; anda heavy chain complementarity determining region (CDR) having an aminoacid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%,89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; and a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibodies comprise (a) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) alight chain complementarity determining region (CDR) having an aminoacid sequence 100% identical to at least one of the amino acid sequencesset forth as SEQ ID NOS: 9-11; and a heavy chain complementaritydetermining region (CDR) having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 12-14.

Methods of Treatment

Provided herein are methods of treating cancer in an individual in needthereof comprising administration of an antibody that selectively bindto a complex comprising a non-classical HLA-I (e.g. HLA-E) and aneoantigen as disclosed herein.

In some embodiments, the antibody selectively binds to a complexcomprising a non-classical HLA-I (e.g. HLA-E) and a neoantigen. In someembodiments, the antibody does not have a binding affinity to thenon-classical HLA-I alone. In some embodiments, the antibody does nothave a binding affinity to the neoantigen alone. In some embodiments,the antibody does not have a binding affinity to a complex comprisingthe non-classical HLA-I and a non-relevant neoantigen.

In some embodiments, the neoantigen is expressed by an antigenprocessing machinery (APM)-proficient cell. In some embodiments, theneoantigen is expressed by a TAP1/2-proficient cell. In someembodiments, the neoantigen comprises, consists essentially of, orconsists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL).

In some embodiments, the non-classical HLA-I is HLA-E, HLA-F, HLA-G, orHLA-H. In some embodiments, the non-classical HLA-I is HLA-E. In someembodiments, the HLA-E is HLA-E*0101. In some embodiments, the HLA-E isHLA-E*0103.

In some embodiments, the antibody selectively binds to the complexcomprising the HLA-E and the neoantigen. In some embodiments, theantibody selectively binds to the complex comprising the HLA-E*0101 andthe neoantigen. In some embodiments, the antibody selectively binds tothe complex comprising the HLA-E*0103 and the neoantigen. In someembodiments, the antibody selectively binds to the complex comprisingthe HLA-E*0101 and the neoantigen, and to the complex of the HLA-E*0103and the neoantigen. In some embodiments, the complex comprises the HLA-Eand a neoantigen according to SEQ ID NO: 18 (VMAPRTLFL).

In some embodiments, the antibody comprises a light chain variabledomain (VL) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 7. In someembodiments, the VL has an amino acid sequence at least about 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence set forthas SEQ ID NO: 7. In some embodiments, the VL has an amino acid sequence100% identical to an amino acid sequence set forth as SEQ ID NO: 7.

In some embodiments, the antibody comprises a heavy chain variabledomain (VH) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 8. In some embodimentsthe VH has an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 8. In some embodiments, the VH has an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 8.

In some embodiments, the antibody comprises a light chain variabledomain (VL) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 7 and a heavy chainvariable domain (VH) having an amino acid sequence at least about 70%identical to an amino acid sequence set forth as SEQ ID NO: 8. In someembodiments the VL has an amino acid sequence at least about 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence set forthas SEQ ID NO: 7 and the VH has an amino acid sequence at least about75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 8. In some embodiments, the VL has an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 7 and the VH has an amino acid sequence 100% identical to an aminoacid sequence set forth as SEQ ID NO: 8.

In some embodiments, the antibody comprises a light chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3. In someembodiments, the antibody comprises a light chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, the antibody comprises a light chainCDR sequence having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3.

In some embodiments, the antibody comprises a heavy chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6. In someembodiments, the antibody comprises a heavy chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the antibody comprises a heavy chainCDR sequence having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6.

In some embodiments, the antibody comprises a light chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3 and a heavychain CDR sequence having an amino acid sequence at least about 70%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the antibody comprises a light chainCDR sequence having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3 and a heavy chain CDR sequencehaving an amino acid sequence at least about 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the antibody comprises alight chain CDR sequence having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 1-3and a heavy chain CDR sequence having an amino acid sequence at leastabout 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a light chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 1, a light chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 2, and a light chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 3.In some embodiments, the antibody comprises at least one of a lightchain a light chain CDR1 having an amino acid sequence at least about75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 1, a light chain CDR2 having an amino acidsequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%,88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identicalto an amino acid sequence set forth as SEQ ID NO: 2, and a light chainCDR3 having an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 3. In some embodiments, the antibody comprises at least one of alight chain CDR1 having an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 1, a light chain CDR2 havingan amino acid sequence 100% identical to an amino acid sequence setforth as SEQ ID NO: 2, and a light chain CDR3 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 3.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a heavy chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 5, a heavy chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 6.In some embodiments, the antibody comprises at least one of a heavychain CDR1 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 4, a heavy chain CDR2 having an amino acid sequence at leastabout 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 5, a heavy chain CDR3 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 6. In someembodiments, the antibody comprises at least one of a heavy chain CDR1having an amino acid sequence 100% identical to an amino acid sequenceset forth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 5, a heavy chain CDR3 having an amino acid sequence 100% identicalto an amino acid sequence set forth as SEQ ID NO: 6.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a light chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 1, a light chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 2, a light chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 3,a heavy chain CDR1 having an amino acid sequence at least about 70%identical to an amino acid sequence set forth as SEQ ID NO: 4, a heavychain CDR2 having an amino acid sequence at least about 70% identical toan amino acid sequence set forth as SEQ ID NO: 5, and a heavy chain CDR3having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 6. In some embodiments, theantibody comprises at least one of a light chain CDR1 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 1, a lightchain CDR2 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 2, a light chain CDR3 having an amino acid sequence at leastabout 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 3, a heavy chain CDR1 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 4, a heavychain CDR2 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 5, and a heavy chain CDR3 having an amino acid sequence atleast about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an aminoacid sequence set forth as SEQ ID NO: 6. In some embodiments, theantibody comprises at least one of a light chain CDR1 having an aminoacid sequence 100% identical to an amino acid sequence set forth as SEQID NO: 1, a light chain CDR2 having an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 2, a lightchain CDR3 having an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 3, a heavy chain CDR1 having an aminoacid sequence 100% identical to an amino acid sequence set forth as SEQID NO: 4, a heavy chain CDR2 having an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 5, and aheavy chain CDR3 having an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 6.

In some embodiments, the antibody is a bispecific antibody. In someembodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 70%identical to an amino acid sequence set forth as SEQ ID NO: 7; or aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; or a heavy chain variable domain (VH) comprising an aminoacid sequence at least 70% identical to an amino acid sequence set forthas SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 7; or a heavy chain variable domain (VH) comprisingan amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence atleast 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 15; or a heavy chain variable domain(VH) comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 7; or a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 8; and (b) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 15; or a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 70%identical to an amino acid sequence set forth as SEQ ID NO: 7; and aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; and a heavy chain variable domain (VH) comprising anamino acid sequence at least 70% identical to an amino acid sequence setforth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 7; and a heavy chain variable domain (VH) comprisingan amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence atleast 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 15; and a heavy chain variable domain(VH) comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 7; and a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 8; and (b) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 15; and a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 70% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 70% identical to at least one of the aminoacid sequences set forth as SEQ ID NOS: 9-11; or a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to at least oneof the amino acid sequences set forth as SEQ ID NOS: 1-3; or a heavychain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; or a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 1-3; or a heavy chain complementarity determining region(CDR) having an amino acid sequence 100% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 9-11; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 70% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 70% identical to at least one of the aminoacid sequences set forth as SEQ ID NOS: 9-11; and a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to at least oneof the amino acid sequences set forth as SEQ ID NOS: 1-3; and a heavychain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; and a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 1-3; and a heavy chain complementarity determining region(CDR) having an amino acid sequence 100% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 9-11; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the antibody is a murine antibody. In someembodiments, the antibody is a chimeric antibody. In some embodiments,the antibody is a camelid antibody. In some embodiments, the antibody isa humanized antibody. In some embodiments, the antibody is a humanantibody.

In some embodiments, the antibody is a TCR-like antibody. In someembodiments, the antibody is a single domain antibody. In someembodiments, the single domain antibody is a camelid single domainantibody.

In some embodiments, the antibody is a multispecific antibody. In someembodiments, the antibody is a bispecific antibody. In some embodiments,the antibody is a bispecific T cell engager (BiTE). In some embodiments,the BiTE binds to a CD3 protein associated with a T cell receptor (TCR).In some embodiments, the BiTE binds to a CD3c protein associated with aT cell receptor (TCR). In some embodiments, the antibody is amultifunctional antibody.

In some embodiments, the antibody further comprises a conjugatedtherapeutic moiety. Therapeutic moiety include, but are not limited to,a cytotoxin, a chemotherapeutic drug, an immunosuppressant, and aradioisotope. A cytotoxin or cytotoxic agent includes any agent that isdetrimental to (e.g., kills) cells. Examples include, but are notlimited to, taxol, cytochalasin B, gramicidin D, ethidium bromide,emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine,colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione,mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone,glucocorticoids, procaine, tetracaine, lidocaine, propranolol, andpuromycin and analogs or homologs thereof. Suitable chemotherapeuticagents include, but are not limited to, anti metabolites (e.g.,methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, fludarabin,5-fluorouracil, decarbazine, hydroxyurea, azathiprin, gemcitabin andcladribin), alkylating agents (e.g., mechlorethamine, thioepa,chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU),cyclophosphamide, busulfan, dibromomannitol, streptozotocin, mitomycinC, and cis-dichlorodiamine platinum (II) (DDP) cisplatin),anthracyclines (e.g., daunorubicin (formerly daunomycin) anddoxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin),bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents(e.g., vincristine, vinblastine, docetaxel, paclitaxel and vinorelbin).Suitable radioisotopes include, but are not limited to, iodine-131,yttrium-90 or indium-Ill. Further examples of therapeutic moieties are aprotein or polypeptide possessing a desired biological activity. Suchproteins may include, for example, an enzymatically active toxin, oractive fragment thereof, such as abrin, ricin A, pseudomonas exotoxin,or diphtheria toxin; a protein such as tumor necrosis factor orinterferon-γ; or biological response modifiers such as, for example,lymphokines, interleukin-1 (IL-1), interleukin-2 (IL-2), interleukin-6(IL-6), granulocyte macrophage colony stimulating factor (GM-CSF),granulocyte colony stimulating factor (G-CSF), or other growth factors.

In some embodiments, the selective binding of the antibody to thecomplex comprising the non-classical HLA-I (e.g. HLA-E) and theneoantigen induces an immune response. In some embodiments, the immuneresponse comprises activation of NK cells. In some embodiments, theselective binding of the antibody to the CD3 protein associated with aTCR induces an immune response. In some embodiments, the immune responsecomprises activation of T cells. In some embodiments, the T cell is aCD8+ T cell. In some embodiments, the immune response comprisesactivation of cytotoxic T cells (CTLs).

In some embodiments, the cancer is breast cancer. In some embodiments,the cancer is kidney cancer. In some embodiments, the cancer is lungcancer. In some embodiments, the cancer is ovarian cancer. In someembodiments, the cancer is colorectal cancer. In some embodiments, thecancer is pancreatic cancer. In some embodiments, the cancer ischoriocarcinoma. In some embodiments, the cancer is non-small cell lungcarcinoma (NSCLC). In some embodiments, the cancer is gastric cancer. Insome embodiments, the cancer is cervical cancer. In some embodiments,the cancer is head and neck cancer. In some embodiments, the cancer is amyeloma. In some embodiments, the cancer is a leukemia. In someembodiments, the cancer is a lymphoma. In some embodiments, the canceris acute myeloid leukemia (AML). In some embodiments, the cancer ismultiple myeloma. In some embodiments, the cancer is a myelodysplasticsyndrome. In some embodiments, the cancer is a B-cell malignancy. Insome embodiments, the cancer is mantel cell lymphoma.

In some embodiments, the cancer cell differentially expresses theneoantigen. In some embodiments, the cancer cell differentiallyexpresses the HLA-E. In some embodiments, the cancer cell differentiallyexpresses the complex comprising the HLA-E and the neoantigen.

Any suitable route of administration is contemplated for use with themethods disclosed herein. In some embodiments, the antibody isadministered by intravenous administration. In some embodiments, theantibody is administered by subcutaneous administration. In someembodiments, the antibody is administered locally. In some embodiments,the antibody is administered systemically (e.g., intravenously,intramuscularly, subcutaneously, intradermally, orally, intranasally,sublingually). In some embodiments, the antibody is formulated as asalve, lotion or emulsion. In some embodiments, the antibody isformulated as a solution. In some embodiments, the antibody isformulated for topical, oral, buccal, or nasal administration.

In some embodiments, the individual is monitored prior to administrationof the antibody. Symptoms are identified and their severity is assessed.An antibody as described herein is administered alone or in combinationwith additional treatments, singly or multiply over time as discussedherein or known to one of skill in the art. In some embodiments, theindividual is monitored such that the efficacy of the treatment regimenis determined. In some embodiments, a treatment regimen is modified inresponse to preliminary treatment outcomes, such that treatment dose orfrequency or dose and frequency is altered so as to attain a desiredlevel of subject response in light of symptom alleviation, side effectreduction, or a combination of symptom alleviation and side effectreduction.

Pharmaceutical Compositions

Also disclosed herein are pharmaceutical compositions comprising (a) anantibody that selectively bind to a complex comprising a non-classicalHLA-I (e.g. HLA-E) and a neoantigen as disclosed herein and (b) apharmaceutically acceptable carrier or excipient.

In some embodiments, the antibody comprises a light chain variabledomain (VL) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 7. In someembodiments, the VL has an amino acid sequence at least about 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence set forthas SEQ ID NO: 7. In some embodiments, the VL has an amino acid sequence100% identical to an amino acid sequence set forth as SEQ ID NO: 7.

In some embodiments, the antibody comprises a heavy chain variabledomain (VH) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 8. In some embodimentsthe VH has an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 8. In some embodiments, the VH has an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 8.

In some embodiments, the antibody comprises a light chain variabledomain (VL) having an amino acid sequence at least about 70% identicalto an amino acid sequence set forth as SEQ ID NO: 7 and a heavy chainvariable domain (VH) having an amino acid sequence at least about 70%identical to an amino acid sequence set forth as SEQ ID NO: 8. In someembodiments the VL has an amino acid sequence at least about 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence set forthas SEQ ID NO: 7 and the VH has an amino acid sequence at least about75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 8. In some embodiments, the VL has an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 7 and the VH has an amino acid sequence 100% identical to an aminoacid sequence set forth as SEQ ID NO: 8.

In some embodiments, the antibody comprises a light chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3. In someembodiments, the antibody comprises a light chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 1-3. In some embodiments, the antibody comprises a light chainCDR sequence having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3.

In some embodiments, the antibody comprises a heavy chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6. In someembodiments, the antibody comprises a heavy chain CDR sequence having anamino acid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the antibody comprises a heavy chainCDR sequence having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 4-6.

In some embodiments, the antibody comprises a light chain CDR sequencehaving an amino acid sequence at least about 70% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 1-3 and a heavychain CDR sequence having an amino acid sequence at least about 70%identical to at least one of the amino acid sequences set forth as SEQID NOS: 4-6. In some embodiments, the antibody comprises a light chainCDR sequence having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to at least one of the amino acidsequences set forth as SEQ ID NOS: 1-3 and a heavy chain CDR sequencehaving an amino acid sequence at least about 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6. In some embodiments, the antibody comprises alight chain CDR sequence having an amino acid sequence 100% identical toat least one of the amino acid sequences set forth as SEQ ID NOS: 1-3and a heavy chain CDR sequence having an amino acid sequence at leastabout 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 4-6.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a light chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 1, a light chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 2, and a light chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 3.In some embodiments, the antibody comprises at least one of a lightchain a light chain CDR1 having an amino acid sequence at least about75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequenceset forth as SEQ ID NO: 1, a light chain CDR2 having an amino acidsequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%,88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identicalto an amino acid sequence set forth as SEQ ID NO: 2, and a light chainCDR3 having an amino acid sequence at least about 75%, 80%, 81%, 82%,83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% identical to an amino acid sequence set forth as SEQ IDNO: 3. In some embodiments, the antibody comprises at least one of alight chain CDR1 having an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 1, a light chain CDR2 havingan amino acid sequence 100% identical to an amino acid sequence setforth as SEQ ID NO: 2, and a light chain CDR3 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 3.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a heavy chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 5, a heavy chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 6.In some embodiments, the antibody comprises at least one of a heavychain CDR1 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 4, a heavy chain CDR2 having an amino acid sequence at leastabout 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 5, a heavy chain CDR3 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 6. In someembodiments, the antibody comprises at least one of a heavy chain CDR1having an amino acid sequence 100% identical to an amino acid sequenceset forth as SEQ ID NO: 4, a heavy chain CDR2 having an amino acidsequence 100% identical to an amino acid sequence set forth as SEQ IDNO: 5, a heavy chain CDR3 having an amino acid sequence 100% identicalto an amino acid sequence set forth as SEQ ID NO: 6.

In some embodiments, the antibody comprises an HLA-E and a neoantigencomprise at least one of a light chain CDR1 having an amino acidsequence at least about 70% identical to an amino acid sequence setforth as SEQ ID NO: 1, a light chain CDR2 having an amino acid sequenceat least about 70% identical to an amino acid sequence set forth as SEQID NO: 2, a light chain CDR3 having an amino acid sequence at leastabout 70% identical to an amino acid sequence set forth as SEQ ID NO: 3,a heavy chain CDR1 having an amino acid sequence at least about 70%identical to an amino acid sequence set forth as SEQ ID NO: 4, a heavychain CDR2 having an amino acid sequence at least about 70% identical toan amino acid sequence set forth as SEQ ID NO: 5, and a heavy chain CDR3having an amino acid sequence at least about 70% identical to an aminoacid sequence set forth as SEQ ID NO: 6. In some embodiments, theantibody comprises at least one of a light chain CDR1 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 1, a lightchain CDR2 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 2, a light chain CDR3 having an amino acid sequence at leastabout 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 3, a heavy chain CDR1 having an aminoacid sequence at least about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 4, a heavychain CDR2 having an amino acid sequence at least about 75%, 80%, 81%,82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, or 99% identical to an amino acid sequence set forth asSEQ ID NO: 5, and a heavy chain CDR3 having an amino acid sequence atleast about 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an aminoacid sequence set forth as SEQ ID NO: 6. In some embodiments, theantibody comprises at least one of a light chain CDR1 having an aminoacid sequence 100% identical to an amino acid sequence set forth as SEQID NO: 1, a light chain CDR2 having an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 2, a lightchain CDR3 having an amino acid sequence 100% identical to an amino acidsequence set forth as SEQ ID NO: 3, a heavy chain CDR1 having an aminoacid sequence 100% identical to an amino acid sequence set forth as SEQID NO: 4, a heavy chain CDR2 having an amino acid sequence 100%identical to an amino acid sequence set forth as SEQ ID NO: 5, and aheavy chain CDR3 having an amino acid sequence 100% identical to anamino acid sequence set forth as SEQ ID NO: 6.

In some embodiments, the antibody is a bispecific antibody. In someembodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 70%identical to an amino acid sequence set forth as SEQ ID NO: 7; or aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; or a heavy chain variable domain (VH) comprising an aminoacid sequence at least 70% identical to an amino acid sequence set forthas SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 7; or a heavy chain variable domain (VH) comprisingan amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence atleast 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 8′7%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 15; or a heavy chain variable domain(VH) comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 7; or a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 8; and (b) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 15; or a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 70%identical to an amino acid sequence set forth as SEQ ID NO: 7; and aheavy chain variable domain (VH) comprising an amino acid sequence atleast 70% identical to an amino acid sequence set forth as SEQ ID NO: 8;and (b) a light chain variable domain (VL) comprising an amino acidsequence at least 70% identical to an amino acid sequence set forth asSEQ ID NO: 15; and a heavy chain variable domain (VH) comprising anamino acid sequence at least 70% identical to an amino acid sequence setforth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence at least 75%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence setforth as SEQ ID NO: 7; and a heavy chain variable domain (VH) comprisingan amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to an amino acid sequence set forth as SEQ ID NO: 8; and (b) alight chain variable domain (VL) comprising an amino acid sequence atleast 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to an amino acidsequence set forth as SEQ ID NO: 15; and a heavy chain variable domain(VH) comprising an amino acid sequence at least 75%, 80%, 81%, 82%, 83%,84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, or 99% identical to an amino acid sequence set forth as SEQ ID NO:16.

In some embodiments, the bispecific antibody comprises (a) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 7; and a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 8; and (b) a light chainvariable domain (VL) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 15; and a heavy chainvariable domain (VH) comprising an amino acid sequence 100% identical toan amino acid sequence set forth as SEQ ID NO: 16.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 70% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 70% identical to at least one of the aminoacid sequences set forth as SEQ ID NOS: 9-11; or a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to at least oneof the amino acid sequences set forth as SEQ ID NOS: 1-3; or a heavychain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; or a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 1-3; or a heavy chain complementarity determining region(CDR) having an amino acid sequence 100% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 9-11; or a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 1-3; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence at least 70% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 70% identical to at least one of the aminoacid sequences set forth as SEQ ID NOS: 9-11; and a heavy chaincomplementarity determining region (CDR) having an amino acid sequenceat least 70% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequenceat least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to at least oneof the amino acid sequences set forth as SEQ ID NOS: 1-3; and a heavychain complementarity determining region (CDR) having an amino acidsequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to atleast one of the amino acid sequences set forth as SEQ ID NOS: 4-6; and(b) a light chain complementarity determining region (CDR) having anamino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%, 85%, 86%,87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%identical to at least one of the amino acid sequences set forth as SEQID NOS: 9-11; and a heavy chain complementarity determining region (CDR)having an amino acid sequence at least 75%, 80%, 81%, 82%, 83%, 84%,85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or99% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 12-14.

In some embodiments, the bispecific antibody comprises (a) a light chaincomplementarity determining region (CDR) having an amino acid sequence100% identical to at least one of the amino acid sequences set forth asSEQ ID NOS: 1-3; and a heavy chain complementarity determining region(CDR) having an amino acid sequence 100% identical to at least one ofthe amino acid sequences set forth as SEQ ID NOS: 4-6; and (b) a lightchain complementarity determining region (CDR) having an amino acidsequence 100% identical to at least one of the amino acid sequences setforth as SEQ ID NOS: 9-11; and a heavy chain complementarity determiningregion (CDR) having an amino acid sequence 100% identical to at leastone of the amino acid sequences set forth as SEQ ID NOS: 12-14.

In some embodiments, excipients for use with the compositions disclosedherein include maleic acid, tartaric acid, lactic acid, citric acid,acetic acid, sodium bicarbonate, sodium phosphate, histidine, glycine,sodium chloride, potassium chloride, calcium chloride, zinc chloride,water, dextrose, N-methylpyrrolidone, dimethyl sulfoxide,N,N-dimethylacetamide, ethanol, propylene glycol, polyethylene glycol,diethylene glycol monoethyl ether, and surfactantpolyoxyethylene-sorbitan monooleate.

In some embodiments, the compositions further comprise an additionaltherapeutic agent. In some embodiments, the therapeutic agent is achemotherapeutic agent. The chemotherapeutic agents can include, amongothers, cytotoxic agents, anti-metabolite agents (e.g., folateantagonists, purine analogs, pyrimidine analogs, etc.), topoisomeraseinhibitors (e.g., camptothecin derivatives, anthracenedione,anthracyclines, epipodophyllotoxins, quinoline alkaloids, etc.),anti-microtubule agents (e.g., taxanes, vinca alkaloids), proteinsynthesis inhibitors (e.g., cephalotaxine, camptothecin derivatives,quinoline alkaloids), alkylating agents (e.g., alkyl sulfonates,ethylenimines, nitrogen mustards, nitrosoureas, platinum derivatives,triazenes, etc.), alkaloids, terpenoids, and kinase inhibitors.

In some embodiments, the antibody and the therapeutic agent are in thesame formulation. In some embodiments, the antibody and the therapeuticagent are in different formulation. In some embodiments, antibodydescribed herein is used prior to the administration of the othertherapeutic agent. In some embodiments, antibody described herein isused concurrently with the administration of the other therapeuticagent. In some embodiments, antibody described herein is used subsequentto the administration of the other therapeutic agent.

Pharmaceutical formulations are made to be compatible with a particularlocal, regional or systemic administration or delivery route. Thus,pharmaceutical formulations include carriers, diluents, or excipientssuitable for administration by particular routes. Specific non-limitingexamples of routes of administration for compositions herein areparenteral, e.g., intravenous, intra-arterial, intradermal,intramuscular, subcutaneous, intra-pleural, transdermal (topical),transmucosal, intra-cranial, intra-spinal, intra-ocular, rectal, oral(alimentary), mucosal administration, and any other formulation suitablefor the treatment method or administration protocol.

Solutions or suspensions used for parenteral application include: asterile diluent such as water for injection, saline solution, fixedoils, polyethylene glycols, glycerine, propylene glycol or othersynthetic solvents; antibacterial agents such as benzyl alcohol ormethyl parabens; antioxidants such as ascorbic acid or sodium bisulfate;chelating agents such as ethylenediaminetetraacetic acid; buffers suchas acetates, citrates or phosphates; and agents for the adjustment oftonicity such as sodium chloride or dextrose. In some embodiments, pH isadjusted with acids or bases, such as hydrochloric acid or sodiumhydroxide.

Pharmaceutical formulations for injection include sterile aqueoussolutions (where water soluble) or dispersions and sterile powders forthe extemporaneous preparation of sterile injectable solutions ordispersion. For intravenous administration, suitable carriers includephysiological saline, bacteriostatic water, Cremophor EL™ (BASF,Parsippany, N.J.), or phosphate buffered saline (PBS). In someembodiments, the carrier is a solvent or dispersion medium containing,for example, water, ethanol, polyol (for example, glycerol, propyleneglycol, and liquid polyetheylene glycol, and the like), or suitablemixtures thereof. Fluidity is maintained, in some embodiments, forexample, by the use of a coating such as lecithin, by the maintenance ofthe required particle size in the case of dispersion, and by the use ofsurfactants. Antibacterial and antifungal agents include, for example,parabens, chlorobutanol, phenol, ascorbic acid, and thimerosal. Isotonicagents, for example, sugars; polyalcohols such as mannitol or sorbitol;or sodium chloride, in some embodiments, are included in thecomposition. In some cases, also included is an agent which delaysabsorption, for example, aluminum monostearate or gelatin prolongsabsorption of injectable compositions.

Sterile injectable formulations are prepared by incorporating the activecomposition in the required amount in an appropriate solvent with one ora combination of above ingredients. Generally, dispersions are preparedby incorporating the active composition into a sterile vehiclecontaining a basic dispersion medium and any other ingredient. In thecase of sterile powders for the preparation of sterile injectablesolutions, methods of preparation include, for example, vacuum dryingand freeze-drying which yields a powder of the active ingredient plusany additional desired ingredient from a previously prepared solutionthereof.

For transmucosal or transdermal administration, penetrants appropriateto the barrier to be permeated are used in the formulation. Suchpenetrants are known in the art, and include, for example, fortransmucosal administration, detergents, bile salts, and fusidic acidderivatives. In some embodiments, transmucosal administration isaccomplished through the use of nasal sprays, inhalation devices (e.g.,aspirators) or suppositories. For transdermal administration, the activecompounds are formulated into ointments, salves, gels, creams orpatches.

In some embodiments, the pharmaceutical formulations are prepared withcarriers that protect against rapid elimination from the body, such as acontrolled release formulation or a time delay material such as glycerylmonostearate or glyceryl stearate. The formulations, in someembodiments, are also delivered using articles of manufacture such asimplants and microencapsulated delivery systems to achieve local,regional or systemic delivery or controlled or sustained release.

EXAMPLES

The following examples are given for the purpose of illustrating variousembodiments of the invention and are not meant to limit the presentinvention in any fashion. The present examples, along with the methodsdescribed herein are presently representative of preferred embodiments,are exemplary, and are not intended as limitations on the scope of theinvention. Changes therein and other uses which are encompassed withinthe spirit of the invention as defined by the scope of the claims willoccur to those skilled in the art.

Example 1. Generation of Antibodies

R4 clone 1 (R4c1) was first isolated from a semi-synthetic humanantibody phage library. In brief, this was performed as follows. A humanscFv antibody phage display library (1.42×10⁹ clones) constructed byCreative Biolabs (HuScL-2 phage library) was used to screen againstHLA-G signal peptide/HLA-E*0103 complex (ABI-V-0025; VMAPRTLFL (SEQ IDNO: 18)). Briefly, 30 μg of a mix of five internal peptide/HLA-A2*0201complexes (ABI-AV-007-Biot, ABI-AV-0010-Biot, ABI-AV-0014-Biot,ABI-AV-0019 and ABI-AB-0030-Biot) were used for depletion andpre-blocking, followed by enrichment with 50 μg of ABI-V-0025. Theenrichment factor of the phage library was 2.73×10⁶ (phage input:5×10¹¹, phage output: 1.83×10⁵). This process was repeated for thesecond round of biopanning, with an enrichment factor of 4.03×10². Thisprocess was repeated for the third round of biopanning, but using 20 μgof the complex mix for depletion (enrichment factor 2.31×10²). For thefourth round of biopanning, 50 μg of ABI-V-0018 was used for depletionand pre-blocking, followed by enrichment with 50 μg of ABI-V-0025(enrichment factor 44). From the phage output of the fourth round ofbiopanning, 40 clones were picked and sequenced, revealing one uniquesequence. This clone was designated as R4 clone 1 (R4c1) (FIG. 1A-FIG.1B). To determine affinity of the R4c1 antibody it was diluted in 0.1%BSA, 0.1% Tween20, PBS dilution/wash buffer and immobilized on theForteBio Octet Protein A biosensor tip at 20 ug/ml for 50 seconds. Tipswere subsequently washed in dilution/wash buffer for 50 seconds.Antibody binding association was measured using six concentrations(serial dilutions starting at 100 ug/ml in dilution/wash buffer) ofcanonical 0025-E 01:03 monomer complex over the course of 150 secondsimmediately followed by dissociation (dilution/wash buffer) for 150seconds (FIG. 1C). The equilibrium dissociation constant (KD) for cloneR4 (clone 1) was determined to be 411 nM (FIG. 1D).

Affinity Maturation of R4c1: Cycle 2:

To increase the binding affinity of clone R4c1 to V-0025, the R4c1 scFvconstruct was cloned into a yeast vector to create a library of R4c1mutants by changing two amino acid in the CDRH3 region. In brief, theyeast display library generated used EBY100 yeast competent cells thatwere prepared for transformation by incubation in 100 mM lithium acetate(LiAc, Sigma) plus 10 mM Dithiothreitol (DTT, Sigma). Electroporationwas achieved using a GenePulser (BioRad) with 60 OD prepared yeast per 2mm cuvette (BioRad) in 10 mM LiAc, with 1 μg of scFv clones and 100 ngof pYES3 (ThermoFisher) altered to include Aga2 after the GAL1 promoterand a Flag (DYKDDDDK) epitope tag (SEQ ID NO: 19) after the scFv. Todetermine the number of transformants, 100 μL of 1/10, 1/100 and 1/1000dilutions of transformed yeast were spread on glucose plates minustryptophan and uracil (D-UT, Teknova). Transformation efficiency wascalculated after incubation at 30° C. for 2 days. Random clones werethen picked for insertion diversity by PCR. Yeast were grown at 30° C.with shaking at 200 rpm in CM broth with glucose minus tryptophan anduracil (D-UT, Teknova). scFv surface induction was achieved by growingyeast for at least 20 hrs in CM broth with galactose minus tryptophanand uracil (G-UT, Teknova) supplemented with 0.1% raffinose (Sigma)(GR-UT).

Yeast were stained after first blocking with 2% BSA in PBS 0.05% tween(blocking buffer) for 30 minutes to 1 hour at room temperature withrotation. Yeast were than incubated with biotinylated peptide/HLAcomplex monomers on ice for 30 minutes to 90 minutes, depending onmonomer concentration. Secondary labeling was achieved with StreptaivdinPE (ThermoFisher) and DYKDDDDK epitope tag (SEQ ID NO: 19) Alexa Fluor488 (R&D Systems) on ice for 30 minutes. Samples were acquired using aLSR II flow cytometer (BD Biosciences) or a CytoFLEX S (BeckmenCoulter). Data was prepared using Flowjo Software version 10 or FACSDiva(BD Biosciences).

Screening was achieved by either magnetic-activated cell sorting (MACS)or fluorescence-activated cell sorting (FACS), using at least ten-foldcoverage of the library size. For MACS screening, yeast were firstblocked with 2% BSA in PBS 0.05% tween (blocking buffer) for 30 minutesto 1 hour at room temperature with rotation. Biotinylated peptide/HLAcomplex monomers were than incubated on ice for 30 minutes at 1 μM.Secondary labeling was achieved using streptavidin microbeads (MiltenyiBiotec). Labeled yeast were poured over a MACS Column on a MACSSeparator (Miltenyi Biotec). Yeast bound to the column were harvestedand considered as an enriched population. For FACS screening, yeast wereprepared as described in the flow cytometry section. Samples were sortedusing a FACS Aria II (BD Bioscience). Sorted yeast were considered anenriched population.

Unique scFv clones were identified by plating. After two days at 30° C.,individual colonies were picked and analyzed by PCR. Full-length humanIgG1 of the selected clones were produced in Expi293F (ThermoFisher)cells. Briefly, antibody variable regions were subcloned into mammalianexpression vectors, with matching human kappa light-chain constantregion (pFUSE2ss-CLIg-hK, Invivogen) and human IgG1 constant region(pFUSEss-CHIg-hG1, Invivogen) sequences. Vectors were transfected at a2:3 light chain:heavy chain ratio. Antibodies were purified with proteinA resin (Genscript) and confirmed in reducing and non-reducingconditions by electrophoresis.

The first round of selection was performed using 1 μM of V-0025. Gatingwas based on binding by the parent (R4c1); parent exhibited 0.11% toV-0025 while the cycle 2 library exhibited 1.66%, indicating thepresence of higher affinity binders (FIG. 2A). Round two sort used 100nM V-0025. The cycle 2 library showed 0.23% binding while the parentshowed 0%; no binding was observed to a negative control (V-0018) (FIG.2B). The third round of sorting used 10 nM V-0025, with 1.05% bindingobserved (FIG. 2C). The final round of sorting incorporated a Koff;briefly, V-0025 was incubated for 30 minutes at 100 nM, followed byincubation with R4c1 in full-length hIgG1 format (R4c1-IgG1). R4c1-IgG1was added at 500 nM for various time points to act as an antibody sink.Thus, a final round of sorting was done using Koff pressure to selectfor top clones (FIG. 2D). To identify unique clones, individual colonieswere picked and sequenced (n=48). Ten unique clones were identified(Table 2) and binding affinity was compared with the parent clone. Ofthe unique clones, six clones were shown to have at least 100-foldgreater binding to V-0025 compared with the parent (FIG. 3A-FIG. 3D).One clone (clone R2A_1), in addition to having increased bindingaffinity, also exhibited superior selectivity and was selected foradditional analysis (FIG. 3E-FIG. 3G).

TABLE 2 Unique clones and sequence modifications Index Name CDRH3 1parent XXXXXXX 2 R2A_1 XLXNXXX 3 R2A_3 XHXTXXX 4 R2A_8 XHXAXXX 5 R2B_1XHXNXXX 6 R2B_3 XHXRXXX 7 R2B_8 XHXWXXX 8 oR2A_1 XYXTXXX 9 oR2A_2XVXXXXX 10 oR2A_3 XLXSXXX 11 R3A2 XHXVXXX

Characterization of Clone R2A_1:

Clone R2A_1 was converted from scFv to full-length hIgG1 for testing byELISA and for cell staining. Based on ELISA data, affinity was measuredat about 20 nM (data not shown), which represents over 10-fold increasein affinity compared with the parent clone, R4c1. To check antibodystability, binding was compared after storing the antibody at 37° C. for72 hr. As shown in FIG. 4 binding by R2A_1 to A549-B4 cells was heavilycompromised when the antibody had been stored at 37° C. for 72 hr.

Stabilization of Clone R2A_1:

In order to increase the stability of R2A_1, several alterations weremade to amino acids in the framework regions of both VH and VL. For theVL, the CDRL2 was modified to match its germline (IGKV1-39). This alsoremoved an N-linked glycosylation site. For the VH, in framework region1, Gln5 was changed to match germline (Leu5). Also in framework region1, Met18 was changed to match germline (Leu18). The stabilized clone wasreferred to as R4c1 H1-L1. The scFv sequence was inserted into yeast fordisplay and compared with the parent (R2A_1). While there was some lossin affinity, there was no alteration in specificity (data not shown).

To generate an antibody with greater affinity and broader specificity, ayeast display library was generated based off of R4c1-H1-L1 andintroduced four amino acid mutations in the CDRL3 region, meaning that44% of the CDRL3 sequence could potentially be altered from the parent.As shown in FIG. 5A (R0), very view clones exhibited binding to target,although there were a select group that had greater binding to targetthan parent. A sort was done (R1 out) using target at 10 nM with gatingmade to only select clones with greater binding affinity than the parent(FIG. 6A). The R1 library was checked for binding to target, and asubstantial increase was observed in the number of clones binding withimproved affinity compared with parent (FIG. 5A, R1). To select forclones with enhanced affinity, the R1 library was subjected to Koffpressure. Briefly, the library was incubated with 10 nM of target. Afterwashing, 1 μM of the parent clone as a full length IgG1 was added for 15to 120 minutes to act as an antibody sink. Any yeast still showingbinding to target indicate a greater Koff compared with the parentclone. As shown in FIG. 5B, yeast clones were observed that maintainedbinding with target even after 120 minutes in the antibody sink Toselect for these highly improved clones, a final sort (R2 out) was doneusing Koff with target at 10 nM and parent clone-IgG1 at 1 μM for 45minutes as the antibody sink, with sort gate made to collect only theclones with the highest affinity (FIG. 6B). After sorting, individualclones were isolated and sequenced (n=60). 12 unique clones were found(Table 3) and were checked for binding to target at 1 and 0.1 nM (FIG.7). The top binding clone to target was selected for additional testing(clone R2-C02) (FIG. 7 black arrow).

TABLE 3 Unique clones Clone CDRL3 % occurrence Parent XXXXXXXXX R2-B04FXXXGXXXQ 6.25 R2-A01 XCXXXXXGL 6.25 R2-C02 XXXXAXXSL 6.25 R2-B02XXXXAXXSY 6.25 R2-A03 XXXXCXXMF 6.25 R2-C03 XXXXSXXQF 6.25 R2-A09XXXXSXXQW 6.25 R2-A07 XXXXXFWEX 12.5 R2-A06 XXXXXXFSP 25 R2-C12XXXXXXXCL 6.25 R2-B12 XXXXXXXQF 6.25 R2-A02 XXXXXXXSL 6.25

Example 2. Characterization of the Top Clone by ELISA, FACS andLabel-Free Technology

The top clone identified from yeast display was made as hIgG1 and testedfor specificity and affinity. R2-C02 exhibited high affinity and thefine specificity (data not shown). R2-C02 was given the designationABX0011 (or ABX-11) and produced in CHO cells using a 1-L transienttransfection protocol followed by purification by Protein-A affinitychromatography. Sample purity was determined by SDS-PAGE reduced (twobands representing heavy and light chains) and non-reduced (single bandat ˜150 kD) (FIG. 8A) and by observing a single peak by size exclusionchromatography (FIG. 8B). ABX0011 was first tested against a broad arrayof peptide/HLA complexes. As shown in FIG. 9A, HLA-E clone 3D12 binds toall HLA-E complexes, including peptide-empty HLA-E. However, ABX0011only recognized the non-classical HLA, HLA-G signal peptide/HLA-Ecomplex and did not recognize closely related signal peptides fromclassical HLA (FIG. 9B). Furthermore, it did not recognize anyadditional peptide/HLA-E complexes, or any peptide/HLA-A2 complexes. Forthe ELISAs run, highly pure HLA-E 01:03 and HLA-A2 02:01 monomercomplexes (diluted in 0.1% BSA, 0.1% Tween20, PBS dilution/wash buffer)were immobilized on neutravidin coated microarray plates at 0.25 ug/mlfor one hour at RT. The plate was subsequently washed in dilution/washbuffer 5×. Antibodies diluted to 1 ug/ml in dilution/wash buffer wereincubated for one hour at RT and plate was subsequently washed indilution/wash buffer 5×. Anti-mouse and anti-human HRP conjugate wasdiluted 1:5000 in dilution/wash buffer and incubated for 30 minutes atRT then subsequently washed 5× in dilution/wash buffer. TMB substratewas added to plate wells and incubated for 15 minutes at 50 ul/well. 1NHCl stop solution was added at 50 μl/well before reading plate at 450 nmabsorbance.

To determine whether ABX0011 recognized classical HLA I signal peptidepresented by two different phenotypes of HLA-E, namely HLA-E 01:03 andHLA-E 01:01, monomer complexes of each were used in an ELISA asdescribed above. No difference was observed between binding to the HLA-Gsignal peptide in the two HLA-E alleles; HLA-E*0101 and *0103 (FIG. 9B).

Affinity Determination for ABX0011:

The binding affinity of ABX0011 was determined using label-freetechnology (Resonant Sensors, Inc., ResoSens instrument). Thedissociation equilibrium constant (KD) was found to be 7.9 nM or animprovement of more than 50-fold over the R4c1 clone (FIG. 1D, FIG. 9C).The method for determining the affinity was as follows. ThermofisherCapture Select™ Biotin anti-IgG-Fc (Hu) (diluted PBS buffer) wasimmobilized on neutravidin coated Bionetic label-free microarray plateat 5 μg/ml until binding reached equilibrium. Plate subsequently waswashed 3× with 0.1% BSA, 0.1% Tween20, PBS dilution/wash buffer. ABX0011(5 μg/ml in dilution/wash buffer) was captured by anti-IgG-Fc untilbinding reached equilibrium. Plate was subsequently washed indilution/wash buffer 3×. V-0025 HLA-E 01:03 monomer complex (serialdilutions starting at 5 μg/ml in dilution/wash buffer) was added towells and allowed to incubate for 20 minutes to determine the bindingassociation and then immediately followed by dissociation (dilution/washbuffer) for approximately 25 minutes. Binding affinity calculated usingTracedrawer kinetic analysis software.

ABX0011 detection sensitivity could be observed by ELISA. In brief,V-0025 HLA-E 01:03 monomer complex (serial dilutions starting at 2 μg/mlin 0.1% BSA, 0.1% Tween20, PBS buffer) was immobilized on neutravidincoated microarray plate for one hour at RT. Plate was subsequentlywashed in dilution/wash buffer 5×. Antibodies were diluted to 1 μg/ml indilution/wash buffer and incubated for one hour at RT and subsequentlywashed in dilution/wash buffer 5×. Anti-mouse and anti-human HRPconjugate was diluted 1:5000 in dilution/wash buffer and incubated for30 minutes at RT and then subsequently washed 5× in dilution/washbuffer. TMB substrate incubated for 15 minutes at 50 ul/well. 1N HClstop solution added at 50 μl/well and plate is read at 450 nm. In FIG.9D, ABX0011 and HLA-E were bound at 1 μg/mL and tested against atitration of the antigen V-0025. Similarly, in FIG. 9E, V-0025 was boundat 0.25 μg/mL and tested against a titration of ABX0011 and HLA-E usinga similar ELISA protocol as described for FIG. 9A. To check stability,ABX0011 was left at 37° C. for 7 to 14 days, then checked againstABX0011 stored at 4° C. ABX0011 was diluted to 1 μg/ml in mouse serumincubated at 37° C. for 7 and 14 days (4° C. control was prepared inserum at time of testing) and added to wells with immobilized HLA-E01:03 monomer complexes and run in an ELISA as described above. As shownin FIG. 9F, binding was not altered, indicating good stability ofABX0011.

To further characterize specificity, K562 cells transfected to expressHLA-E were first peptide-pulsed with the V-0025 peptide, then tested forbinding to HLA-E and ABX0011. As shown in FIG. 9G, V-0025 peptide pulsedcells showed loading, based on an increase in HLA-E expression comparedwith no peptide (unpulsed). ABX0011 showed specificity for HLA-G loadedon HLA-E, with no binding to unpulsed cells (FIG. 9G). K562 cellstransfected to express HLA-E were next pulsed with all of the classicalHLA signal peptides, the non-classical HLA signal peptide, and two otherHLA-E binding peptides namely, V-0038 and P-0550. All peptide pulsedcells showed loading, based on an increase in HLA-E expression comparedwith no peptide (unpulsed) (FIG. 9H). Again, ABX0011 (as exemplified byABX0012) recognized only the V-0025 peptide in K562 cells expressingHLA-E (FIG. 9I). Background staining was approximately 500 antibodymolecules/cell. To further assess the fine binding specificity ofABX0011, K562 cells expressing HLA-E were peptide pulsed with 20 μg/mlof either V-0025, V-0034, V-0044, V-0046, P-0550 or left unpulsed andthen stained with ABX0011 at various concentrations ranging from 10 to<0.001 μg/ml. Again, ABX0011 showed selectivity only for the V-0025peptide with only minimal detection of two closely related peptides,V-0044 and V-0046 when used at a concentration of 10 ug/ml (FIG. 9J).

An alanine scan based on the peptide sequence for V-0025 (HLA-G signalpeptide) indicated that the binding of ABX0011 to peptide/HLA-E complexis mainly dependent on the amino acid at position 8 with less dependencethough still critical contact at amino acid positions 2, 7 and 9 (FIG.9K). Overall, these results indicate that ABX0011 exhibits selectivityto non-classical HLA-G peptide signal sequence, with high affinity andstability.

Example 3. ABX0011 Binds with Selectivity to Target Positive Cancer CellLines

To confirm specificity, binding of ABX0011 was tested in a variety oftumor cell lines, human peripheral blood mononuclear cells (huPBMC) andprimary human umbilical vascular endothelial cells (huVEC). Since HLA-Eis known to be highly expressed in normal placenta trophoblasts, thechoriocarcinoma trophoblastic cell line JEG-3 was used as a positivecontrol for binding by both HLA-E and ABX0011 (FIG. 10A). To confirm therestriction of ABX0011 to HLA-E/peptide complexes, binding was tested inHLA-E-deficient JEG-3 (JEG-3 Eko). As shown in FIG. 10A, middle panel,HLA-E and ABX0011 were unable to bind to JEG-3 Eko cells. To confirmthat the HLA-G signal peptide is dependent on Tap1/2 expression, ABX0011was used to stain Tap-1 knock-out JEG3 cells (JEG-3 Tap-1ko). As shownin FIG. 10A, lower panel, there is still some expression of HLA-Ewithout Tap-1, however, ABX0011 exhibits no binding, indicating that itonly recognizes the HLA-G signal peptide processed via the conventionalpathway. Specificity was similarly shown using huPBMC and huVEC cells.Human PBMCs, in particular B-cells and NK cells express high levels ofHLA-E (FIG. 10B). In contrast, healthy huPBMC do not express HLA-G andtherefore huPBMC lack binding with ABX0011 (FIG. 10B). Additionally,huVEC after O/N treatment with IFN-γ express high levels of HLA-E.However, ABX0011 (as exemplified by ABX0010) does not bind to huVECs inthe absence or presence of IFN-γ treatment (FIG. 10C).

HLA-E is known to be increased in multiple types of cancers. Therefore,several cancer cell lines were analyzed for HLA-E expression and ABX0011binding. Since HLA-E is upregulated when cells are stressed orstimulated, binding was observed in both unstimulated andIFNγ-stimulated cells. FIG. 11A illustrates two cell lines (THP-1 andOCI-AML3) that express HLA-E and HLA-G and bind ABX0011 withoutstimulation, and one cell line (KG-1) that express HLA-E and not HLA-Gand do not bind ABX0011. Further, binding to AML cell lines with ABX0011(as exemplified by ABX-0012) using a range of concentrations (10 to<0.001 μg/ml) revealed good selectivity for ABX0011 (FIG. 11B). Theseresults indicate that ABX0011 may be used to target AML cancers as wellas other cancers that express both HLA-E and HLA-G. K562 cellsexpressing HLA-E and mutated JEG3 cells were acquired from (Thorbald vanHall; Leiden University Medical Center). In some instances, cellsrequired stimulation overnight with IFNγ (10 ng/ml) to induce HLA-Eexpression.

For all cell binding studies (with exception ABX0012, the ABX0011 withmouse IgG1), cells were blocked with anti-human CD16 (clone KD1),anti-human CD32 (clone AT10), and anti-human CD64 (clone 10.1) (Bio-Rad)or with Human TruStain FcX (BioLegend) at room temperature for 5-10 minbefore surface staining with ABX0011 or hIgG1 (clone QA16A12, BioLegend)at 1 μg/mL or with AF647-labeled ABX0011 or hIgG1, and with HLA-E in PEor PE-Cy7 (clone 3D12, BioLegend) at 4° C. for 30 min in 100 μL. Cellswere then washed with staining buffer (PBS with 2 mM EDTA) followed bysecondary staining cocktail including Goat-anti-human/AF647 (JacksonImmunoResearch) and zombie/aqua viability stain (BioLegend). Cells wereincubated at 4° C. for 30 min in 100 μL. Cells were washed twice withstaining buffer and fixed using fluorofix (BioLegend). Samples wereacquired using an LSR II flow cytometer (BD Biosciences) or a CytoFLEX S(Beckmen Coulter). Data was prepared using Flowjo Software version 10(BD Biosciences).

Example 4. ABX0011 Induces NK Mediated Antibody-Dependent CellularCytotoxicity (ADCC) of Target Cells

The primary objective of these studies was to determine the ability ofABX0011 and to induce NK cell mediated ADCC of target cells. To test theability of ABX0011 to induce ADCC, primary human NK cells wereco-cultured with various tumor target cells in the presence of ABX0011.Blood from healthy volunteers was acquired from Carter BloodCare. PBMCswere isolated by density-gradient centrifugation using Ficoll-Paque PLUS(GE Healthcare) or Lymphoprep (StemCell Technologies). Natural Killer(NK) cells were purified using EasySep human NK cell enrichment kit(StemCell Technologies).

As shown in FIG. 12A, after four-hour co-culture of NK cells withpeptide-pulsed K562.E cells, ABX0011 was able to significantly increasekilling of K562.E cells in a dose-dependent manner compared to isotypecontrol (FIG. 12A) and upregulate NK CD107a expression (FIG. 12B) andmore specifically induce an increase of CD107a expression on NKG2A+ NKcells (FIG. 12C). Similar results were also found when using THP-1 cells(FIG. 12D). These results indicate that ABX0011 can be used to promotetumor cytotoxicity by inducing NK cell ADCC activity.

Example 5. Creating a Bispecific Antibody Capable of Retargeting T-Cellsto Lyse Tumor Target Cells

ABX0030 was constructed as a bispecific antibody to be able to activateCD3+ T-cells to kill tumor cells. ABX0030 was made as an Fab-Fc-scFvconstruct containing ABX0011 (Fab) coupled to a single-chain (scFv) ofclone SP34 (mouse anti-human CD3e) on an non-glycosylated (N297Dmutation) human IgG1 scaffold. A_20 amino acid linker(GKPGSGKPGSGKPGSGKPGS (SEQ ID NO: 20)) was used to covalently join theABX0011 Fab with the SP34 scFv. ABX0030 was produced in a transientlytransfected CHO cell and purified on a Protein-A affinity column.ABX0030 was run on a reducing SDS-PAGE gel and shows the expected twobands at the anticipated MW (25 kD for the ABX0011 light chain and ˜55kD for ABX0030 without light chain) (FIG. 13A). Greater than 91% ofABX0030 migrates as a single peak on an analytical SEC-HPLC column (FIG.13B).

The binding affinity of ABX0030 was determined using a label-free assay(ResoSens instrument, Resonant Sensors, Inc). Briefly, V-0025 HLA-E01:03 monomer complex (diluted in 0.1% BSA, 0.1% Tween20, PBS buffer)was immobilized on neutravidin coated Bionetic label-free microarrayplate at 5 μg/ml until binding reached equilibration. Plate wassubsequently washed in dilution/wash buffer 3×. The ABX0030 (serialdilutions starting at 10 μg/ml in dilution/wash buffer) bindingassociation was evaluated for approximately 20 minutes and immediatelyfollowed by dissociation (dilution/wash buffer) for approximately 15minutes. Binding affinity calculated using Tracedrawer kinetic analysissoftware. The findings revealed a dissociation equilibrium constant of8.97 nM (FIG. 14).

Binding properties of ABX0030: ABX0030 binding to target tumor cells wasfirst confirmed. As shown in FIG. 15, ABX0030 binds to V-0025 peptidepulsed K562 cells expressing HLA-E similarly to the parent clone(ABX0011). This binding is HLA-E restricted, as HLA-E negative parentK562 cells do not exhibit binding to ABX0030 (FIG. 15). Binding to Tcells in peripheral blood mononuclear cells (PBMCs) was used to confirmthe anti-CD3 arm (FIG. 16). The K562 cell line was acquired from ATCC.K562 cells expressing HLA-E cells were acquired from (Dr. Thorbald vanHall, Leiden University Medical Center, Netherlands). Cells werecultured per the supplier's recommendation. For cell binding, cells wereblocked with anti-human CD16 (clone KD1), anti-human CD32 (clone AT10),and anti-human CD64 (clone 10.1) (Bio-Rad) or with Human TruStain FcX(BioLegend) at room temperature for 5-10 min before surface stainingwith unlabeled antibody at 1 μg/mL or with AF647-labeled antibody, andwith HLA-E/PE-Cy7 (clone 3D12, BioLegend) at 4° C. for 30 min in 50-100μL. Cells were then washed with staining buffer (PBS with 2 mM EDTA)followed by secondary staining cocktail including Goat-anti-human orGoat-anti-mouse/AF647 (if using unlabeled antibody, JacksonImmunoResearch) and zombie/aqua viability stain (BioLegend). Cells wereincubated at 4° C. for 30 min in 50 μL. Cells were washed twice withstaining buffer and fixed using fluorofix (BioLegend). Samples wereacquired using a CytoFLEX S (Beckmen Coulter). Data was prepared usingFlowJo Software version 10 (BD Biosciences).

ABX0030 Enhances Target-Specific Cytotoxicity of Primary CTLs:

To assess the activity of ABX0030, primary human CTLs were co-culturedwith either K562 (HLA-E positive) or K562.E pulsed with V-0025 peptideand tested with a titration of ABX0030. As shown in FIG. 17A, ABX0030induced dose-dependent cytotoxicity in a target-specific manner;unpulsed K562.E cells were not affected. Further, ABX0030 does notinduce expression of CD25 (48 hrs) (FIG. 17B) and IFNγ (48 hrs) (FIG.17C) in CD8+ T cells cultured with target negative K562.E cells.

These results were confirmed using THP-1 cells (ABX0011 binding datashown in FIG. 11A). After 48 hr co-culture, ABX030 induced cytotoxicityin a dose-dependent manner (FIG. 18A). ABX0030 was also able to induceCD8+ T cell activation, as shown by the dose-dependent induction ofCD25, CD107a, perforin and IFNγ (FIG. 18B-FIG. 18E). These resultsindicate that ABX0030 enhances lysis of tumor cells by CD8+ T cells in atarget-specific manner.

To perform in vitro T-cell activation assays, blood from healthyvolunteers was acquired from Carter BloodCare (Arlington, Tex.). PBMCswere isolated by density-gradient centrifugation using Ficoll-Paque PLUS(GE Healthcare) or Lymphoprep (StemCell Technologies). CD8+ T cells werepurified using EasySep human CD8+ T cell enrichment kit (StemCellTechnologies). The enriched human CD8+ T cells (CTLs) were restedovernight at 1×10⁶/mL. Target cells were labeled with CFSE (ThemoFisher)and added to the rested CTLs at a ratio of 10:1 (E:T). CD3/CD28stimulation (ImmunoCult human CD3/CD28 T cell activator, StemCellTechnologies) was used as a positive control. In the last 4 hr ofculture, monensin and anti-human CD107a/BV421 (BioLegend) was added toall wells. Cells were harvested after 24 to 48 hrs. Cells were stainedwith anti-human CD8/APC (BioLegend), anti-human CD25/PE-Cy7 (Tonbo) andZombie/Aqua viability marker (BioLegend). For intracellular staining,cells were fixed and permeabilized using BD cytofix/cytoperm solution(BD) and were stained with anti-human Perforin/PerCP-Cy5.5 (BioLEgend)and anti-human IFNγ/PE-Dazzle 594 (BioLegend). Samples were acquiredusing a CytoFLEX S (Beckmen Coulter). Data was prepared using FlowJoSoftware version 10 (BD Biosciences).

TABLE 4 List of peptide/HLA complexes Peptide Peptide SEQ ID HLA familyID Sequence NO: complex Gene Origin Classical V-0021 VMAPRTLIL 21 HLA-EHLA-C HLA-Cw*01, -Cw*03, HLA -Cw*04, -Cw*05, -Cw*06, -Cw*08, -Cw*12,-Cw*14, -Cw*16 and -Cw*17:02 V-0034 VMAPRTLVL 22 HLA-E HLA-A*02, -A*23,-A*24, -A*25, -A*26, -A*34:02, -A*34:06, -A*43, -A*66 and -A*69 V-0035VTAPRTLLL 23 HLA-E HLA-B*13, -B*18, -B*27, -B*37, -B*40, -B*44, -B*47,-B*54, B*55 -B*56, -B*59, -B*82 and B*83 V-0037 IMAPRTLVL 24 HLA-EHLA-A*34:01 V-0040 VMAPQALLL 25 HLA-E HLA-Cw*17:01, Cw*17:03 and-Cw*17:05 V-0041 VMAPRALLL 26 HLA-E HLA-Cw*06:17, -Cw*07 and -Cw*18V-0042 VMAPRTLLL 27 HLA-E HLA-A*01, -A*03, -A*11, -A*29, -A*30, A*31,-A*32, -A*33, -A*36, and A*74, HLA-Cw02 and Cw*15 V-0043 VMAPRTLTL 28HLA-E HLA-Cw*08:09 V-0044 VMAPRTVLL 29 HLA-E HLA-B*07, -B*08, -B*14,-B*38, -B*39, -B*42, -B*48, -B*67, -B*73 and -B*81 V-0045 VMPPRTLLL 30HLA-E HLA-A*80 V-0046 VTAPRTVLL 31 HLA-E HLA-B*15, -B35, -B*40, -B*41,-B*44:18, -B*45, -B*46 -B*49, -B*50, -B*51, -B*52, -B*53, -B*57, -B*58and -B*78 Non-classical V-0025 VMAPRTLFL 18 HLA-E HLA-G HLA AdditionalV-0038 QMRPVSRVL 32 HLA-E Hsp60 P-0550 RAARLPPLL 33 HLA-E PODXL2

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments described herein may beemployed. It is intended that the following claims define the scope ofthe invention and that methods and structures within the scope of theseclaims and their equivalents be covered thereby.

What is claimed is:
 1. A monoclonal antibody, or an antigen-bindingfragment thereof, that selectively binds to a complex comprising anHLA-E and a peptide that comprises, consists essentially of, or consistsof a sequence according to SEQ ID NO: 18 (VMAPRTLFL), comprising: (i) alight chain CDR1 having the amino acid sequence set forth as SEQ ID NO:1, a light chain CDR2 having the amino acid sequence set forth as SEQ IDNO: 2, a light chain CDR3 having the amino acid sequence set forth asSEQ ID NO: 3; and (ii) a heavy chain CDR1 having the amino acid sequenceset forth as SEQ ID NO: 4, a heavy chain CDR2 having the amino acidsequence set forth as SEQ ID NO: 5, and a heavy chain CDR3 having theamino acid sequence set forth as SEQ ID NO:
 6. 2. The monoclonalantibody of claim 1, wherein the monoclonal antibody or antigen-bindingfragment thereof does not have a binding affinity to (i) the HLA-Ealone; or (ii) the peptide alone.
 3. The monoclonal antibody of claim 2,wherein the peptide consists of a sequence according to SEQ ID NO: 18(VMAPRTLFL).
 4. A bispecific T cell engager, comprising the monoclonalantibody of claim 1 or an antigen binding-fragment thereof.
 5. Thebispecific T cell engager of claim 4, wherein the bispecific T cellengager binds to a CD3 protein associated with a T cell receptor.
 6. Amonoclonal antibody, or an antigen-binding fragment thereof, thatselectively binds to a complex comprising an HLA-E and a peptide thatcomprises, consists essentially of, or consists of a sequence accordingto SEQ ID NO: 18 (VMAPRTLFL), comprising: (i) a light chain variabledomain (VL) comprising the amino acid sequence set forth as SEQ ID NO:7; and (ii) a heavy chain variable domain (VH) comprising the amino acidsequence set forth as SEQ ID NO:
 8. 7. The monoclonal antibody of claim6, wherein the monoclonal antibody or antigen-binding fragment thereofdoes not have a binding affinity to (i) the HLA-E alone; or (ii) thepeptide alone.
 8. The monoclonal antibody of claim 7, wherein thepeptide consists of a sequence according to SEQ ID NO: 18 (VMAPRTLFL).9. A bispecific T cell engager, comprising the monoclonal antibody ofclaim 6 or an antigen-binding fragment thereof.
 10. The bispecific Tcell engager of claim 9, wherein the bispecific T cell engager binds toa CD3 protein associated with a T cell receptor.
 11. A method attreating a cancer that expresses both HLA-E and HLA-G in an individualin need thereof, comprising administering to the individual an effectiveamount of a monoclonal antibody of claim 1 or an antigen-bindingfragment thereof, to thereby treat the cancer.
 12. The method of claim11, wherein the cancer is breast cancer, kidney cancer, lung cancer,ovarian cancer, colorectal cancer, pancreatic cancer, choriocarcinoma,non-small cell lung carcinoma (NSCLC), gastric cancer, cervical cancer,head and neck cancer, leukemia, lymphoma, myeloma, multiple myeloma(MM), acute myeloid leukemia (AML), myelodysplastic syndrome, a B-cellmalignancy, or mantel cell lymphoma.
 13. A method of treating a cancerthat expresses both HLA-E and HLA-G in an individual in need thereof,comprising administering to the individual an effective amount of amonoclonal antibody of claim 6 or an antigen-binding fragment thereof,to thereby treat the cancer.